TY  - CONF
AU  - Gligorijević, Nikola
AU  - Stanić-Vučinić, Dragana
AU  - Mutić, Tamara
AU  - Lujić, Tamara
AU  - Ćirković Veličković, Tanja
PY  - 2023
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/6524
AB  - Microplastic represents one of the major types of pollutants in modern era. Over several years of research in the field of microplastic, there are still many unknown gaps, including the effects and mechanisms of action of these particles on human health. Studies in this field conducted experiments on cells and human tissues or animals like rats and mice. While these studies suggest the toxic effects of microplastic, it is not clear if concentrations used for exposure are relevant for humans. Also, most of the studies used spherical polystyrene, which does not reflect well the diversity of microplastic particles found in nature. Another gap is lack of studies describing direct interactions of microplastics and proteins. While it is generally known that proteins form corona around microplastic particles, affinity studies and consequences on protein structure are usually missing. The aim of this work was to analyze interaction of a major egg white protein and allergen, ovalbumin to several to microplastic particles, including polystyrene (PS) of 120 and 500 μm in size and polyethylene terephthalate (PET) of 120 μm in size. Binding affinity was determined at both acidic, pH 3 and neutral, pH 7 conditions, at the room temperature, by measuring bulk ovalbumin concentration in supernatants at the equilibrium time. Several binding models, including Langmuir,
Freundlich, Redlich–Peterson and Guggenheim-Anderson-de Boer (GAB), were used to determine binding parameters. The formation of soft and hard corona was analyzed according to the published protocol [1]. Structural analysis was performed using near and far-UV CD spectrometry.
Obtained results showed that ovalbumin binds to both PS and PET. All binding models indicated that ovalbumin binds with higher affinity to tested microplastics on pH 3, compared to pH 7, with the highest affinity being calculated for PS 120 μm. Further analysis showed that ovalbumin forms both soft and hard corona onto the surface of all three microplastics. Structural alterations of ovalbumin as a consequence of its interaction with microplastic was shown to be both pH and
microplastic type dependent. Also, more pronounced effect on its tertiary structure was observed, compared to secondary. At pH3, tertiary structure of bulk ovalbumin becomes destabilized, especially in the presence of PET 120 μm and PS 500 μm, while at pH 7, structural stabilization is observed, especially in the presence of PS 120 μm. Considering that the microplastic was discovered in eggs [2], obtained results suggest that direct interactions of native ovalbumin with microplastic particles could have influence on its structure and thus affect its techno-functional properties.
PB  - Belgrade : Serbian Chemical Society
C3  - Book of Abstracts of the XXII EuroFoodChem Congress, Belgrade, Serbia, 14-16 June 2023
T1  - Binding and corona formation of ovalbumin to polystyrene and polyethylene terephthalate microplastics under neutral and acidic conditions
SP  - 137
EP  - 137
UR  - https://hdl.handle.net/21.15107/rcub_cer_6524
ER  - 

@conference{
author = "Gligorijević, Nikola and Stanić-Vučinić, Dragana and Mutić, Tamara and Lujić, Tamara and Ćirković Veličković, Tanja",
year = "2023",
abstract = "Microplastic represents one of the major types of pollutants in modern era. Over several years of research in the field of microplastic, there are still many unknown gaps, including the effects and mechanisms of action of these particles on human health. Studies in this field conducted experiments on cells and human tissues or animals like rats and mice. While these studies suggest the toxic effects of microplastic, it is not clear if concentrations used for exposure are relevant for humans. Also, most of the studies used spherical polystyrene, which does not reflect well the diversity of microplastic particles found in nature. Another gap is lack of studies describing direct interactions of microplastics and proteins. While it is generally known that proteins form corona around microplastic particles, affinity studies and consequences on protein structure are usually missing. The aim of this work was to analyze interaction of a major egg white protein and allergen, ovalbumin to several to microplastic particles, including polystyrene (PS) of 120 and 500 μm in size and polyethylene terephthalate (PET) of 120 μm in size. Binding affinity was determined at both acidic, pH 3 and neutral, pH 7 conditions, at the room temperature, by measuring bulk ovalbumin concentration in supernatants at the equilibrium time. Several binding models, including Langmuir,
Freundlich, Redlich–Peterson and Guggenheim-Anderson-de Boer (GAB), were used to determine binding parameters. The formation of soft and hard corona was analyzed according to the published protocol [1]. Structural analysis was performed using near and far-UV CD spectrometry.
Obtained results showed that ovalbumin binds to both PS and PET. All binding models indicated that ovalbumin binds with higher affinity to tested microplastics on pH 3, compared to pH 7, with the highest affinity being calculated for PS 120 μm. Further analysis showed that ovalbumin forms both soft and hard corona onto the surface of all three microplastics. Structural alterations of ovalbumin as a consequence of its interaction with microplastic was shown to be both pH and
microplastic type dependent. Also, more pronounced effect on its tertiary structure was observed, compared to secondary. At pH3, tertiary structure of bulk ovalbumin becomes destabilized, especially in the presence of PET 120 μm and PS 500 μm, while at pH 7, structural stabilization is observed, especially in the presence of PS 120 μm. Considering that the microplastic was discovered in eggs [2], obtained results suggest that direct interactions of native ovalbumin with microplastic particles could have influence on its structure and thus affect its techno-functional properties.",
publisher = "Belgrade : Serbian Chemical Society",
journal = "Book of Abstracts of the XXII EuroFoodChem Congress, Belgrade, Serbia, 14-16 June 2023",
title = "Binding and corona formation of ovalbumin to polystyrene and polyethylene terephthalate microplastics under neutral and acidic conditions",
pages = "137-137",
url = "https://hdl.handle.net/21.15107/rcub_cer_6524"
}

Gligorijević, N., Stanić-Vučinić, D., Mutić, T., Lujić, T.,& Ćirković Veličković, T.. (2023). Binding and corona formation of ovalbumin to polystyrene and polyethylene terephthalate microplastics under neutral and acidic conditions. in Book of Abstracts of the XXII EuroFoodChem Congress, Belgrade, Serbia, 14-16 June 2023
Belgrade : Serbian Chemical Society., 137-137.
https://hdl.handle.net/21.15107/rcub_cer_6524

Gligorijević N, Stanić-Vučinić D, Mutić T, Lujić T, Ćirković Veličković T. Binding and corona formation of ovalbumin to polystyrene and polyethylene terephthalate microplastics under neutral and acidic conditions. in Book of Abstracts of the XXII EuroFoodChem Congress, Belgrade, Serbia, 14-16 June 2023. 2023;:137-137.
https://hdl.handle.net/21.15107/rcub_cer_6524 .

Gligorijević, Nikola, Stanić-Vučinić, Dragana, Mutić, Tamara, Lujić, Tamara, Ćirković Veličković, Tanja, "Binding and corona formation of ovalbumin to polystyrene and polyethylene terephthalate microplastics under neutral and acidic conditions" in Book of Abstracts of the XXII EuroFoodChem Congress, Belgrade, Serbia, 14-16 June 2023 (2023):137-137,
https://hdl.handle.net/21.15107/rcub_cer_6524 .

TY  - CONF
AU  - Lujić, Tamara
AU  - Gligorijević, Nikola
AU  - Stanić-Vučinić, Dragana
AU  - Ćirković Veličković, Tanja
PY  - 2023
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/6525
AB  - Ovalbumin (OVA) is the most abundant protein in chicken egg white. It is one of the major allergens in eggs. Micro- and nanoplatic particles (MNPs) are a widespread contaminant and have been found in food and water. It is still unclear how MNPs might affect human health. However, due to their large surface area they have been found to bind various biopolymers,
including proteins. These biopolymers can be bound more strongly or loosely, and are referred to as hard and soft corona, respectfully [1]. MPs have been found in eggs, in the size range of 50-100 μm [2]. It is shown that these particles can interact with proteins and induce structural changes, but there is still not enough information on this topic [3]. These structural changes could lead to a decreased digestibility in the gastrointestinal tract, which could increase the immune
response to known allergens. The aim of this study was to determine whether there are structural changes present in the OVA after incubation with two types of MPs – 120 μm polyethylene terephthalate (PET) and 120 μm polystyrene (PS) and whether they could influence
digestion of OVA with gastrointestinal enzymes. 20 mg of MPs were incubated with 1.3 mg/mL ovalbumin for 4 h at room temperature in a 20 mM phosphate buffer at pH 7. Bulk ovalbumin was separated from the MPs by centrifugation and by filtration through a 0.22 μm PVDF filter. Soft corona was obtained by washing the MPs with water, and the MPs were later removed as described with bulk ovalbumin. Formation of amyloids was monitored with a Thioflavin T (ThT) assay at room temperature and after thermal treatment, and additional structural analysis was performed by circular dichroism (CD) spectrometry in the far-UV region. Thermal stability was also determined by spectrofluorimetry. Digestion with two proteases (pepsin and trypsin) was performed to determine whether there is a change in the gastrointestinal digestibility of OVA.
Results from the ThT assay show that at room temperature there is no significant difference between the fluorescence emission obtained for all samples, with bulk OVA from both MPs showing a slight decrease. However, there is an increase of fluorescence after thermal treatment in all OVA samples, where OVA from the soft corona emits significantly less fluorescence
than control and bulk samples for both types of MPs. Additionally, soft coronas have been shown to have more β-sheet content than other samples, which is more pronounced for OVA incubated with PET. For the heated samples there is a sharp change from α-helix to β-sheets in all the samples, but it is the most dramatic in the soft coronas. This could impose rigidity to the tertiary structure, which would explain why the ThT molecule does not bind as strongly. Despite differences in both the secondary and tertiary structure, the thermal stability is almost the same in all samples. Digestion of the samples shows that the soft corona incubated with PS tends to be more resistant to trypsin than other samples after 2 min, but it is not significant. For digestion with pepsin there is no difference between the samples. In conjunction with the previous results, which indicates a structural stabilisation of the soft corona at pH 7, it is not surprising that there is an increased resistance to trypsin, compared to pepsin which is a gastric enzyme and for which digestion is performed at an acidic pH. In conclusion, there is a structural change present in samples upon contact with MPs, particularly in the soft corona, of which the most pronounced is a decrease of α-helix content and increase in β-sheet content as determined by far-UV CD.
This leads to a structural stabilization which could further impact the digestibility of the OVA protein and impact its allergenicity. However, this must be confirmed with further experiments.
PB  - Belgrade : Serbian Chemical Society
C3  - Book of Abstracts of the XXII EuroFoodChem Congress, Belgrade, Serbia, 14-16 June 2023
T1  - Investigation of structural changes in ovalbumin induced by two types of MPs and its impact on protein digestibility
SP  - 153
EP  - 153
UR  - https://hdl.handle.net/21.15107/rcub_cer_6525
ER  - 

@conference{
author = "Lujić, Tamara and Gligorijević, Nikola and Stanić-Vučinić, Dragana and Ćirković Veličković, Tanja",
year = "2023",
abstract = "Ovalbumin (OVA) is the most abundant protein in chicken egg white. It is one of the major allergens in eggs. Micro- and nanoplatic particles (MNPs) are a widespread contaminant and have been found in food and water. It is still unclear how MNPs might affect human health. However, due to their large surface area they have been found to bind various biopolymers,
including proteins. These biopolymers can be bound more strongly or loosely, and are referred to as hard and soft corona, respectfully [1]. MPs have been found in eggs, in the size range of 50-100 μm [2]. It is shown that these particles can interact with proteins and induce structural changes, but there is still not enough information on this topic [3]. These structural changes could lead to a decreased digestibility in the gastrointestinal tract, which could increase the immune
response to known allergens. The aim of this study was to determine whether there are structural changes present in the OVA after incubation with two types of MPs – 120 μm polyethylene terephthalate (PET) and 120 μm polystyrene (PS) and whether they could influence
digestion of OVA with gastrointestinal enzymes. 20 mg of MPs were incubated with 1.3 mg/mL ovalbumin for 4 h at room temperature in a 20 mM phosphate buffer at pH 7. Bulk ovalbumin was separated from the MPs by centrifugation and by filtration through a 0.22 μm PVDF filter. Soft corona was obtained by washing the MPs with water, and the MPs were later removed as described with bulk ovalbumin. Formation of amyloids was monitored with a Thioflavin T (ThT) assay at room temperature and after thermal treatment, and additional structural analysis was performed by circular dichroism (CD) spectrometry in the far-UV region. Thermal stability was also determined by spectrofluorimetry. Digestion with two proteases (pepsin and trypsin) was performed to determine whether there is a change in the gastrointestinal digestibility of OVA.
Results from the ThT assay show that at room temperature there is no significant difference between the fluorescence emission obtained for all samples, with bulk OVA from both MPs showing a slight decrease. However, there is an increase of fluorescence after thermal treatment in all OVA samples, where OVA from the soft corona emits significantly less fluorescence
than control and bulk samples for both types of MPs. Additionally, soft coronas have been shown to have more β-sheet content than other samples, which is more pronounced for OVA incubated with PET. For the heated samples there is a sharp change from α-helix to β-sheets in all the samples, but it is the most dramatic in the soft coronas. This could impose rigidity to the tertiary structure, which would explain why the ThT molecule does not bind as strongly. Despite differences in both the secondary and tertiary structure, the thermal stability is almost the same in all samples. Digestion of the samples shows that the soft corona incubated with PS tends to be more resistant to trypsin than other samples after 2 min, but it is not significant. For digestion with pepsin there is no difference between the samples. In conjunction with the previous results, which indicates a structural stabilisation of the soft corona at pH 7, it is not surprising that there is an increased resistance to trypsin, compared to pepsin which is a gastric enzyme and for which digestion is performed at an acidic pH. In conclusion, there is a structural change present in samples upon contact with MPs, particularly in the soft corona, of which the most pronounced is a decrease of α-helix content and increase in β-sheet content as determined by far-UV CD.
This leads to a structural stabilization which could further impact the digestibility of the OVA protein and impact its allergenicity. However, this must be confirmed with further experiments.",
publisher = "Belgrade : Serbian Chemical Society",
journal = "Book of Abstracts of the XXII EuroFoodChem Congress, Belgrade, Serbia, 14-16 June 2023",
title = "Investigation of structural changes in ovalbumin induced by two types of MPs and its impact on protein digestibility",
pages = "153-153",
url = "https://hdl.handle.net/21.15107/rcub_cer_6525"
}

Lujić, T., Gligorijević, N., Stanić-Vučinić, D.,& Ćirković Veličković, T.. (2023). Investigation of structural changes in ovalbumin induced by two types of MPs and its impact on protein digestibility. in Book of Abstracts of the XXII EuroFoodChem Congress, Belgrade, Serbia, 14-16 June 2023
Belgrade : Serbian Chemical Society., 153-153.
https://hdl.handle.net/21.15107/rcub_cer_6525

Lujić T, Gligorijević N, Stanić-Vučinić D, Ćirković Veličković T. Investigation of structural changes in ovalbumin induced by two types of MPs and its impact on protein digestibility. in Book of Abstracts of the XXII EuroFoodChem Congress, Belgrade, Serbia, 14-16 June 2023. 2023;:153-153.
https://hdl.handle.net/21.15107/rcub_cer_6525 .

Lujić, Tamara, Gligorijević, Nikola, Stanić-Vučinić, Dragana, Ćirković Veličković, Tanja, "Investigation of structural changes in ovalbumin induced by two types of MPs and its impact on protein digestibility" in Book of Abstracts of the XXII EuroFoodChem Congress, Belgrade, Serbia, 14-16 June 2023 (2023):153-153,
https://hdl.handle.net/21.15107/rcub_cer_6525 .

TY  - CONF
AU  - Radomirović, Mirjana
AU  - Gligorijević, Nikola
AU  - Stanić-Vučinić, Dragana
AU  - Rajković, Andreja
AU  - Ćirković Veličković, Tanja
PY  - 2023
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/6668
AB  - Tropomyosin has been recognized as one of the most common allergens among shellfish
allergens. Sensitive and specific quantification of traces of allergens present in food
samples is of critical importance for people with food allergies. This study thus aimed to
develop a highly sensitive immuno-PCR method for detecting crustacean tropomyosin in
foods. Method couples conventional sandwich ELISA assay with real-time PCR
amplification of marker DNA. Monoclonal mouse anti-tropomyosin antibody was used as
a capture antibody, while polyclonal rabbit anti-tropomyosin antibody served as a
detection antibody in sandwich ELISA. A double-stranded amino-DNA molecule of 77
base pairs was covalently conjugated to a secondary goat anti-rabbit antibody and
subsequently amplified and quantified by real-time PCR. Tropomyosin was quantified
using highly purified natural shrimp tropomyosin as standard. The sensitivity of immuno-
PCR for quantification of tropomyosin was increased by up to 20-fold compared to
ELISA, demonstrating a ccuracy a s l ow a s 1 9.8 p g/mL. Recovery of tropomyosin in
vegetable soup as a food matrix was in the 87.7–115.6% range, with relative standard
deviations in the 5–24.5% range. Tropomyosin was also quantified in the commercially
available food products. Developed immuno-PCR technique thus shows the potential to be
a method of choice for specific and ultrasensitive detection of tropomyosin in food
samples, with the final aim of reducing risks of accidental food contamination.
PB  - Serbian Biochemical Society
C3  - Serbian Biochemical Society Twelfth Conference, International scientific meeting, “Biochemistry in Biotechnology,” September 21-23, 2023, Belgrade, Serbia
T1  - Immuno-PCR for crustacean tropomyosin quantification
SP  - 130
EP  - 130
UR  - https://hdl.handle.net/21.15107/rcub_cer_6668
ER  - 

@conference{
author = "Radomirović, Mirjana and Gligorijević, Nikola and Stanić-Vučinić, Dragana and Rajković, Andreja and Ćirković Veličković, Tanja",
year = "2023",
abstract = "Tropomyosin has been recognized as one of the most common allergens among shellfish
allergens. Sensitive and specific quantification of traces of allergens present in food
samples is of critical importance for people with food allergies. This study thus aimed to
develop a highly sensitive immuno-PCR method for detecting crustacean tropomyosin in
foods. Method couples conventional sandwich ELISA assay with real-time PCR
amplification of marker DNA. Monoclonal mouse anti-tropomyosin antibody was used as
a capture antibody, while polyclonal rabbit anti-tropomyosin antibody served as a
detection antibody in sandwich ELISA. A double-stranded amino-DNA molecule of 77
base pairs was covalently conjugated to a secondary goat anti-rabbit antibody and
subsequently amplified and quantified by real-time PCR. Tropomyosin was quantified
using highly purified natural shrimp tropomyosin as standard. The sensitivity of immuno-
PCR for quantification of tropomyosin was increased by up to 20-fold compared to
ELISA, demonstrating a ccuracy a s l ow a s 1 9.8 p g/mL. Recovery of tropomyosin in
vegetable soup as a food matrix was in the 87.7–115.6% range, with relative standard
deviations in the 5–24.5% range. Tropomyosin was also quantified in the commercially
available food products. Developed immuno-PCR technique thus shows the potential to be
a method of choice for specific and ultrasensitive detection of tropomyosin in food
samples, with the final aim of reducing risks of accidental food contamination.",
publisher = "Serbian Biochemical Society",
journal = "Serbian Biochemical Society Twelfth Conference, International scientific meeting, “Biochemistry in Biotechnology,” September 21-23, 2023, Belgrade, Serbia",
title = "Immuno-PCR for crustacean tropomyosin quantification",
pages = "130-130",
url = "https://hdl.handle.net/21.15107/rcub_cer_6668"
}

Radomirović, M., Gligorijević, N., Stanić-Vučinić, D., Rajković, A.,& Ćirković Veličković, T.. (2023). Immuno-PCR for crustacean tropomyosin quantification. in Serbian Biochemical Society Twelfth Conference, International scientific meeting, “Biochemistry in Biotechnology,” September 21-23, 2023, Belgrade, Serbia
Serbian Biochemical Society., 130-130.
https://hdl.handle.net/21.15107/rcub_cer_6668

Radomirović M, Gligorijević N, Stanić-Vučinić D, Rajković A, Ćirković Veličković T. Immuno-PCR for crustacean tropomyosin quantification. in Serbian Biochemical Society Twelfth Conference, International scientific meeting, “Biochemistry in Biotechnology,” September 21-23, 2023, Belgrade, Serbia. 2023;:130-130.
https://hdl.handle.net/21.15107/rcub_cer_6668 .

Radomirović, Mirjana, Gligorijević, Nikola, Stanić-Vučinić, Dragana, Rajković, Andreja, Ćirković Veličković, Tanja, "Immuno-PCR for crustacean tropomyosin quantification" in Serbian Biochemical Society Twelfth Conference, International scientific meeting, “Biochemistry in Biotechnology,” September 21-23, 2023, Belgrade, Serbia (2023):130-130,
https://hdl.handle.net/21.15107/rcub_cer_6668 .

TY  - CONF
AU  - Lujić, Tamara
AU  - Gligorijević, Nikola
AU  - Stanić-Vučinić, Dragana
AU  - Bićanin, Maša
AU  - Ćirković Veličković, Tanja
PY  - 2023
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/6651
AB  - Mircoplastics (MPs) are an abundant contaminant in the environment with ingestion being the most common way of exposure for humans. Binding of protein to MPs is proposed to be multilayered with the formation of a soft and hard corona. It has been proven that MPs interact with enzymes present in the digestive system and impact their activity. The aim of this study is to investigate the impact of MPs on the activity of trypsin in simulated intestinal fluid (SIF). For this purpose, two sizes of polypropylene (large – 180-500 μm, small – 63-180 μm) and one size of polyethylene terephthalate (<80 μm) have been studied. Activity in bulk and soft corona was determined in SIF at 405 nm with N-α- Benzoyl-DL-arginine 4-nitroanilide hydrochloride after different times of incubation. Activity in hard corona was determined after 1 h of incubation with the MPs. Although specific activity in the control decreases through time, there is a tendency for all MPs to preserve activity in bulk and soft corona trypsin after 4 h of incubation. Trypsin remains active in the hard corona, with the activity being an order of magnitude lower than in the control, possibly due to significant changes in structure.
PB  - Serbian Biochemical Society
C3  - Serbian Biochemical Society Twelfth Conference, International scientific meeting, “Biochemistry in Biotechnology,” September 21-23, 2023, Belgrade, Serbia
T1  - Impact of MPs on trypsin activity in simulated intestinal fluid
SP  - 145
EP  - 145
UR  - https://hdl.handle.net/21.15107/rcub_cer_6651
ER  - 

@conference{
author = "Lujić, Tamara and Gligorijević, Nikola and Stanić-Vučinić, Dragana and Bićanin, Maša and Ćirković Veličković, Tanja",
year = "2023",
abstract = "Mircoplastics (MPs) are an abundant contaminant in the environment with ingestion being the most common way of exposure for humans. Binding of protein to MPs is proposed to be multilayered with the formation of a soft and hard corona. It has been proven that MPs interact with enzymes present in the digestive system and impact their activity. The aim of this study is to investigate the impact of MPs on the activity of trypsin in simulated intestinal fluid (SIF). For this purpose, two sizes of polypropylene (large – 180-500 μm, small – 63-180 μm) and one size of polyethylene terephthalate (<80 μm) have been studied. Activity in bulk and soft corona was determined in SIF at 405 nm with N-α- Benzoyl-DL-arginine 4-nitroanilide hydrochloride after different times of incubation. Activity in hard corona was determined after 1 h of incubation with the MPs. Although specific activity in the control decreases through time, there is a tendency for all MPs to preserve activity in bulk and soft corona trypsin after 4 h of incubation. Trypsin remains active in the hard corona, with the activity being an order of magnitude lower than in the control, possibly due to significant changes in structure.",
publisher = "Serbian Biochemical Society",
journal = "Serbian Biochemical Society Twelfth Conference, International scientific meeting, “Biochemistry in Biotechnology,” September 21-23, 2023, Belgrade, Serbia",
title = "Impact of MPs on trypsin activity in simulated intestinal fluid",
pages = "145-145",
url = "https://hdl.handle.net/21.15107/rcub_cer_6651"
}

Lujić, T., Gligorijević, N., Stanić-Vučinić, D., Bićanin, M.,& Ćirković Veličković, T.. (2023). Impact of MPs on trypsin activity in simulated intestinal fluid. in Serbian Biochemical Society Twelfth Conference, International scientific meeting, “Biochemistry in Biotechnology,” September 21-23, 2023, Belgrade, Serbia
Serbian Biochemical Society., 145-145.
https://hdl.handle.net/21.15107/rcub_cer_6651

Lujić T, Gligorijević N, Stanić-Vučinić D, Bićanin M, Ćirković Veličković T. Impact of MPs on trypsin activity in simulated intestinal fluid. in Serbian Biochemical Society Twelfth Conference, International scientific meeting, “Biochemistry in Biotechnology,” September 21-23, 2023, Belgrade, Serbia. 2023;:145-145.
https://hdl.handle.net/21.15107/rcub_cer_6651 .

Lujić, Tamara, Gligorijević, Nikola, Stanić-Vučinić, Dragana, Bićanin, Maša, Ćirković Veličković, Tanja, "Impact of MPs on trypsin activity in simulated intestinal fluid" in Serbian Biochemical Society Twelfth Conference, International scientific meeting, “Biochemistry in Biotechnology,” September 21-23, 2023, Belgrade, Serbia (2023):145-145,
https://hdl.handle.net/21.15107/rcub_cer_6651 .

TY  - JOUR
AU  - Radomirović, Mirjana
AU  - Gligorijević, Nikola
AU  - Stanić-Vučinić, Dragana
AU  - Rajković, Andreja
AU  - Ćirković Veličković, Tanja
PY  - 2023
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/6782
AB  - Tropomyosin is the major and predominant allergen among shellfish. This study developed
an ultrasensitive immuno-PCR method for the quantification of crustacean tropomyosin in foods.
The method couples sandwich ELISA with the real-time PCR (rtPCR) amplification of marker DNAs.
Monoclonal anti-TPM antibody was the capture antibody, polyclonal rabbit anti-shrimp tropomyosin
antibody was the detection antibody, while natural shrimp tropomyosin served as the standard. A
double-stranded amino-DNA was covalently conjugated to a secondary anti-rabbit antibody and
subsequently amplified and quantified via rtPCR. The quantification sensitivity of immuno-PCR
was 20-fold higher than analogous ELISA, with LOQ 19.8 pg/mL. The developed immuno-PCR
method is highly specific for the detection of crustacean tropomyosin and is highly precise in a
broad concentration range. Tropomyosin recovery in the spiked vegetable soup was 87.7–115.6%.
Crustacean tropomyosin was also quantified in commercial food products. The reported immuno-
PCR assay is the most sensitive method for the quantification of crustacean tropomyosin and is the
first immuno-PCR-based assay for the quantification of food allergen and food protein in general.
The described method could be easily adapted for the specific and ultrasensitive immuno-PCR-based
detection of traces of any food allergen that is currently being quantified with ELISA, which is of
critical importance for people with food allergies.
PB  - MDPI
T2  - International Journal of Molecular Sciences
T1  - Ultrasensitive Quantification of Crustacean Tropomyosin by Immuno-PCR
VL  - 24
IS  - 20
SP  - 15410
DO  - 10.3390/ijms242015410
ER  - 

@article{
author = "Radomirović, Mirjana and Gligorijević, Nikola and Stanić-Vučinić, Dragana and Rajković, Andreja and Ćirković Veličković, Tanja",
year = "2023",
abstract = "Tropomyosin is the major and predominant allergen among shellfish. This study developed
an ultrasensitive immuno-PCR method for the quantification of crustacean tropomyosin in foods.
The method couples sandwich ELISA with the real-time PCR (rtPCR) amplification of marker DNAs.
Monoclonal anti-TPM antibody was the capture antibody, polyclonal rabbit anti-shrimp tropomyosin
antibody was the detection antibody, while natural shrimp tropomyosin served as the standard. A
double-stranded amino-DNA was covalently conjugated to a secondary anti-rabbit antibody and
subsequently amplified and quantified via rtPCR. The quantification sensitivity of immuno-PCR
was 20-fold higher than analogous ELISA, with LOQ 19.8 pg/mL. The developed immuno-PCR
method is highly specific for the detection of crustacean tropomyosin and is highly precise in a
broad concentration range. Tropomyosin recovery in the spiked vegetable soup was 87.7–115.6%.
Crustacean tropomyosin was also quantified in commercial food products. The reported immuno-
PCR assay is the most sensitive method for the quantification of crustacean tropomyosin and is the
first immuno-PCR-based assay for the quantification of food allergen and food protein in general.
The described method could be easily adapted for the specific and ultrasensitive immuno-PCR-based
detection of traces of any food allergen that is currently being quantified with ELISA, which is of
critical importance for people with food allergies.",
publisher = "MDPI",
journal = "International Journal of Molecular Sciences",
title = "Ultrasensitive Quantification of Crustacean Tropomyosin by Immuno-PCR",
volume = "24",
number = "20",
pages = "15410",
doi = "10.3390/ijms242015410"
}

Radomirović, M., Gligorijević, N., Stanić-Vučinić, D., Rajković, A.,& Ćirković Veličković, T.. (2023). Ultrasensitive Quantification of Crustacean Tropomyosin by Immuno-PCR. in International Journal of Molecular Sciences
MDPI., 24(20), 15410.
https://doi.org/10.3390/ijms242015410

Radomirović M, Gligorijević N, Stanić-Vučinić D, Rajković A, Ćirković Veličković T. Ultrasensitive Quantification of Crustacean Tropomyosin by Immuno-PCR. in International Journal of Molecular Sciences. 2023;24(20):15410.
doi:10.3390/ijms242015410 .

Radomirović, Mirjana, Gligorijević, Nikola, Stanić-Vučinić, Dragana, Rajković, Andreja, Ćirković Veličković, Tanja, "Ultrasensitive Quantification of Crustacean Tropomyosin by Immuno-PCR" in International Journal of Molecular Sciences, 24, no. 20 (2023):15410,
https://doi.org/10.3390/ijms242015410 . .

TY  - CONF
AU  - Gligorijević, Nikola
AU  - Lujić, Tamara
AU  - Mutić, Tamara
AU  - Vasović, Tamara
AU  - de Guzman, Maria Krishna
AU  - Aćimović, Jelena
AU  - Stanić-Vučinić, Dragana
AU  - Ćirković Veličković, Tanja
PY  - 2023
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/6898
AB  - Ovalbumin (OVA), a main protein of egg white, has characteristic structural fold of a
serpin-family of proteins, propensity to fibril formation and stability to digestion.
Microplastics (MPs) contaminating our food can interact with food proteins in the food
matrix and during digestion. In this study adsorption of OVA to polystyrene (PS) (110 μm
and 260 μm), polyethylene terephthalate (PET) (140 μm) MPs were investigated in acidic
(pH 3) and neutral (pH 7) conditions. Formations of corona on MPs were investigated
using isolated OVA and egg white protein extract comparatively. OVA adsorption depends
on MPs size, polymer chemistry and pH, being highest in acidic pH and higher for PS.
Adsorption of OVA to PS and PET reaches dynamic equilibrium after 4h resulting in
disruption of tertiary structure and formation of hard and soft corona around MPs. Shorter
fragments of OVA populate hard corona, while soft corona exclusively consist of full
length OVA, albeit in its non-native conformation. The conformational changes resemble
those induced by heat treatment with re-arrangement of α-β secondary structures.
Structural changes are striking for the OVA in corona around MPs. Soft corona OVA
preserves thermal and proteolytic stability, but loses ability to form fibrils upon heating.
OVA is abundantly present in corona around MPs also in the presence of other egg white
proteins. MPs contaminating food may bind and change structure and functional properties
of main egg white protein.
PB  - Kragujevac, Srbija : Prirodno-matematički fakultet, Univerzitet u Kragujevcu /  Kragujevac, Serbia : Faculty of Science, University of Kragujevac
C3  - VI Simpozijum Srpskog udruženja za proteomiku (SePA) “Razvoj i primena novih metoda proteomike,” Zbornik apstrakata, 2. jun 2023. godine, Kragujevac / VI Symposium of a Serbian proteomic society: „Discussion and Application of  New Methods of Proteomics,“ Book of abstratcs,
T1  - Biocorona formation of hen egg white proteins onto the surface of polystyrene and polyethylene terephthalate
SP  - OP10
EP  - OP10
UR  - https://hdl.handle.net/21.15107/rcub_cer_6898
ER  - 

@conference{
author = "Gligorijević, Nikola and Lujić, Tamara and Mutić, Tamara and Vasović, Tamara and de Guzman, Maria Krishna and Aćimović, Jelena and Stanić-Vučinić, Dragana and Ćirković Veličković, Tanja",
year = "2023",
abstract = "Ovalbumin (OVA), a main protein of egg white, has characteristic structural fold of a
serpin-family of proteins, propensity to fibril formation and stability to digestion.
Microplastics (MPs) contaminating our food can interact with food proteins in the food
matrix and during digestion. In this study adsorption of OVA to polystyrene (PS) (110 μm
and 260 μm), polyethylene terephthalate (PET) (140 μm) MPs were investigated in acidic
(pH 3) and neutral (pH 7) conditions. Formations of corona on MPs were investigated
using isolated OVA and egg white protein extract comparatively. OVA adsorption depends
on MPs size, polymer chemistry and pH, being highest in acidic pH and higher for PS.
Adsorption of OVA to PS and PET reaches dynamic equilibrium after 4h resulting in
disruption of tertiary structure and formation of hard and soft corona around MPs. Shorter
fragments of OVA populate hard corona, while soft corona exclusively consist of full
length OVA, albeit in its non-native conformation. The conformational changes resemble
those induced by heat treatment with re-arrangement of α-β secondary structures.
Structural changes are striking for the OVA in corona around MPs. Soft corona OVA
preserves thermal and proteolytic stability, but loses ability to form fibrils upon heating.
OVA is abundantly present in corona around MPs also in the presence of other egg white
proteins. MPs contaminating food may bind and change structure and functional properties
of main egg white protein.",
publisher = "Kragujevac, Srbija : Prirodno-matematički fakultet, Univerzitet u Kragujevcu /  Kragujevac, Serbia : Faculty of Science, University of Kragujevac",
journal = "VI Simpozijum Srpskog udruženja za proteomiku (SePA) “Razvoj i primena novih metoda proteomike,” Zbornik apstrakata, 2. jun 2023. godine, Kragujevac / VI Symposium of a Serbian proteomic society: „Discussion and Application of  New Methods of Proteomics,“ Book of abstratcs,",
title = "Biocorona formation of hen egg white proteins onto the surface of polystyrene and polyethylene terephthalate",
pages = "OP10-OP10",
url = "https://hdl.handle.net/21.15107/rcub_cer_6898"
}

Gligorijević, N., Lujić, T., Mutić, T., Vasović, T., de Guzman, M. K., Aćimović, J., Stanić-Vučinić, D.,& Ćirković Veličković, T.. (2023). Biocorona formation of hen egg white proteins onto the surface of polystyrene and polyethylene terephthalate. in VI Simpozijum Srpskog udruženja za proteomiku (SePA) “Razvoj i primena novih metoda proteomike,” Zbornik apstrakata, 2. jun 2023. godine, Kragujevac / VI Symposium of a Serbian proteomic society: „Discussion and Application of  New Methods of Proteomics,“ Book of abstratcs,
Kragujevac, Srbija : Prirodno-matematički fakultet, Univerzitet u Kragujevcu /  Kragujevac, Serbia : Faculty of Science, University of Kragujevac., OP10-OP10.
https://hdl.handle.net/21.15107/rcub_cer_6898

Gligorijević N, Lujić T, Mutić T, Vasović T, de Guzman MK, Aćimović J, Stanić-Vučinić D, Ćirković Veličković T. Biocorona formation of hen egg white proteins onto the surface of polystyrene and polyethylene terephthalate. in VI Simpozijum Srpskog udruženja za proteomiku (SePA) “Razvoj i primena novih metoda proteomike,” Zbornik apstrakata, 2. jun 2023. godine, Kragujevac / VI Symposium of a Serbian proteomic society: „Discussion and Application of  New Methods of Proteomics,“ Book of abstratcs,. 2023;:OP10-OP10.
https://hdl.handle.net/21.15107/rcub_cer_6898 .

Gligorijević, Nikola, Lujić, Tamara, Mutić, Tamara, Vasović, Tamara, de Guzman, Maria Krishna, Aćimović, Jelena, Stanić-Vučinić, Dragana, Ćirković Veličković, Tanja, "Biocorona formation of hen egg white proteins onto the surface of polystyrene and polyethylene terephthalate" in VI Simpozijum Srpskog udruženja za proteomiku (SePA) “Razvoj i primena novih metoda proteomike,” Zbornik apstrakata, 2. jun 2023. godine, Kragujevac / VI Symposium of a Serbian proteomic society: „Discussion and Application of  New Methods of Proteomics,“ Book of abstratcs, (2023):OP10-OP10,
https://hdl.handle.net/21.15107/rcub_cer_6898 .

TY  - CONF
AU  - Lujić, Tamara
AU  - Gligorijević, Nikola
AU  - Jovanović, Vesna B.
AU  - Aćimović, Jelena
AU  - Mitić, Dragana
AU  - Vasović, Tamara
AU  - Stojadinović, Marija
AU  - Stanić-Vučinić, Dragana
AU  - Ćirković-Veličković, Tanja
PY  - 2023
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/7473
AB  - Microplastics is abundant in the environment, food and beverages and get ingested by humans.
Its complex interplay with proteins lead to formation of corona. Tightly bound proteins represent
hard corona, while weaker binding partners are found in soft corona. Separation of hard and soft
corona of allergenic proteins of shrimps, eggs and cow’s milk, tropomyosin (TPM), ovalbumin
(OVA) and beta-lactoglobulin (BLG) and identification of binding partners by proteomics was
aim of our study.
Allergenic proteins were purified from egg white, shrimps and cow’s milk. Binding to
polyethylene terephthalate microplastics (PET) (70-100 m) was probed at pH 7 for purified
allergens and egg white proteins. After establishment of binding equilibrium, soft and hard
corona were separated and analyzed by SDS PAGE, followed by identification of bound
proteins by nanoLC-HRMS. Binding of all allergenic proteins was observed in both soft and
hard corona. Soft corona contains exclusively intact, full length OVA, TPM and BLG. Hard
corona is enriched for truncated OVA and oligomers of TPM. OVA fragments are partially or
fully enfolded and have higher level of exposed hydrophobic patches resulting in higher affinity
for PET microplastics. In comparison to OVA and TPM, hard corona of BLG is less abundant
under similar conditions. BLG is compact globular protein with lower level of exposed
hydrophobic patches in comparison to ovalbumin and tropomyosin. In hard corona, trace
amounts of contaminating alfa-lactalbumin become enriched. In the presence of egg white
protein extract OVA forms both SC and HC on microplastics, being the dominant protein of
hard corona (with ovotransferrin). Lysozyme and ovomucin are present only in hard corona.
Both proteins are known for their strong bioactivity and represent a small fraction of total egg
white proteins.
Our results show that allergenic proteins form hard corona on PET microplastics. Among egg
white proteins, minor proteins such as lysozyme and ovomucin become enriched. Denaturing
effect of strong binding to microplastics may change functional characteristics of allergens and
bioactive proteins of foods and should be further investigated in functional assays.
PB  - Italian Proteomics Association
C3  - XVII International Italian Proteomics Association Annual Meeting in partnership with the Hellenic Proteomics Society and Serbian Proteomics Association, "Proteomics and Metabolomics towards Global Health", November 29th-December 1st, 2023, Ospedale Isola Tiberina - Gemelli Isola,  Roma, Italy
T1  - Proteomic insight into allergenic food corona on polyethylene terephthalate microplastics
SP  - 11
EP  - 11
UR  - https://hdl.handle.net/21.15107/rcub_cer_7473
ER  - 

@conference{
author = "Lujić, Tamara and Gligorijević, Nikola and Jovanović, Vesna B. and Aćimović, Jelena and Mitić, Dragana and Vasović, Tamara and Stojadinović, Marija and Stanić-Vučinić, Dragana and Ćirković-Veličković, Tanja",
year = "2023",
abstract = "Microplastics is abundant in the environment, food and beverages and get ingested by humans.
Its complex interplay with proteins lead to formation of corona. Tightly bound proteins represent
hard corona, while weaker binding partners are found in soft corona. Separation of hard and soft
corona of allergenic proteins of shrimps, eggs and cow’s milk, tropomyosin (TPM), ovalbumin
(OVA) and beta-lactoglobulin (BLG) and identification of binding partners by proteomics was
aim of our study.
Allergenic proteins were purified from egg white, shrimps and cow’s milk. Binding to
polyethylene terephthalate microplastics (PET) (70-100 m) was probed at pH 7 for purified
allergens and egg white proteins. After establishment of binding equilibrium, soft and hard
corona were separated and analyzed by SDS PAGE, followed by identification of bound
proteins by nanoLC-HRMS. Binding of all allergenic proteins was observed in both soft and
hard corona. Soft corona contains exclusively intact, full length OVA, TPM and BLG. Hard
corona is enriched for truncated OVA and oligomers of TPM. OVA fragments are partially or
fully enfolded and have higher level of exposed hydrophobic patches resulting in higher affinity
for PET microplastics. In comparison to OVA and TPM, hard corona of BLG is less abundant
under similar conditions. BLG is compact globular protein with lower level of exposed
hydrophobic patches in comparison to ovalbumin and tropomyosin. In hard corona, trace
amounts of contaminating alfa-lactalbumin become enriched. In the presence of egg white
protein extract OVA forms both SC and HC on microplastics, being the dominant protein of
hard corona (with ovotransferrin). Lysozyme and ovomucin are present only in hard corona.
Both proteins are known for their strong bioactivity and represent a small fraction of total egg
white proteins.
Our results show that allergenic proteins form hard corona on PET microplastics. Among egg
white proteins, minor proteins such as lysozyme and ovomucin become enriched. Denaturing
effect of strong binding to microplastics may change functional characteristics of allergens and
bioactive proteins of foods and should be further investigated in functional assays.",
publisher = "Italian Proteomics Association",
journal = "XVII International Italian Proteomics Association Annual Meeting in partnership with the Hellenic Proteomics Society and Serbian Proteomics Association, "Proteomics and Metabolomics towards Global Health", November 29th-December 1st, 2023, Ospedale Isola Tiberina - Gemelli Isola,  Roma, Italy",
title = "Proteomic insight into allergenic food corona on polyethylene terephthalate microplastics",
pages = "11-11",
url = "https://hdl.handle.net/21.15107/rcub_cer_7473"
}

Lujić, T., Gligorijević, N., Jovanović, V. B., Aćimović, J., Mitić, D., Vasović, T., Stojadinović, M., Stanić-Vučinić, D.,& Ćirković-Veličković, T.. (2023). Proteomic insight into allergenic food corona on polyethylene terephthalate microplastics. in XVII International Italian Proteomics Association Annual Meeting in partnership with the Hellenic Proteomics Society and Serbian Proteomics Association, "Proteomics and Metabolomics towards Global Health", November 29th-December 1st, 2023, Ospedale Isola Tiberina - Gemelli Isola,  Roma, Italy
Italian Proteomics Association., 11-11.
https://hdl.handle.net/21.15107/rcub_cer_7473

Lujić T, Gligorijević N, Jovanović VB, Aćimović J, Mitić D, Vasović T, Stojadinović M, Stanić-Vučinić D, Ćirković-Veličković T. Proteomic insight into allergenic food corona on polyethylene terephthalate microplastics. in XVII International Italian Proteomics Association Annual Meeting in partnership with the Hellenic Proteomics Society and Serbian Proteomics Association, "Proteomics and Metabolomics towards Global Health", November 29th-December 1st, 2023, Ospedale Isola Tiberina - Gemelli Isola,  Roma, Italy. 2023;:11-11.
https://hdl.handle.net/21.15107/rcub_cer_7473 .

Lujić, Tamara, Gligorijević, Nikola, Jovanović, Vesna B., Aćimović, Jelena, Mitić, Dragana, Vasović, Tamara, Stojadinović, Marija, Stanić-Vučinić, Dragana, Ćirković-Veličković, Tanja, "Proteomic insight into allergenic food corona on polyethylene terephthalate microplastics" in XVII International Italian Proteomics Association Annual Meeting in partnership with the Hellenic Proteomics Society and Serbian Proteomics Association, "Proteomics and Metabolomics towards Global Health", November 29th-December 1st, 2023, Ospedale Isola Tiberina - Gemelli Isola,  Roma, Italy (2023):11-11,
https://hdl.handle.net/21.15107/rcub_cer_7473 .

TY  - JOUR
AU  - de Guzman, Maria Krishna
AU  - Stanić-Vučinić, Dragana
AU  - Gligorijević, Nikola
AU  - Wimmer, Lukas
AU  - Gasparyan, Manvel
AU  - Lujić, Tamara
AU  - Vasović, Tamara
AU  - Dailey, Lea Ann
AU  - Van Haute, Sam
AU  - Ćirković-Veličković, Tanja
PY  - 2023
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/6466
AB  - Human ingestion of microplastics (MPs) is common and inevitable due to the widespread contamination of food items, but implications on the gastric digestion of food proteins are still unknown. In this study, the interactions between pepsin and polystyrene (PS) MPs were evaluated by investigating enzyme activity and conformation in a simulated human gastric environment in the presence or absence of PS MPs. The impact on food digestion was also assessed by monitoring the kinetics of protein hydrolysis through static in vitro gastric digestion of cow's milk contaminated with PS. The binding of pepsin to PS showed that the surface chemistry of MPs dictates binding affinity. The key contributor to pepsin adsorption seems to be π−π interactions between the aromatic residues and the PS phenyl rings. During quick exposure (10 min) of pepsin to increasing concentrations (222, 2219, 22188 particles/mL) of 10 μm PS (PS10) and 100 μm PS (PS100), total enzymatic activities were not affected remarkably. However, upon prolonged exposure at 1 and 2 h, preferential binding of pepsin to the small, low zeta-potential PS caused structural changes in the protein which led to a significant reduction of its activity. Digestion of cow's milk mixed with PS10 resulted in transient accumulation of larger peptides (10–35 kDa) and reduced bioavailability of short peptides (2–9 kDa) in the gastric phase. This, however, was only observed at extremely high PS10 concentration (0.3 mg/mL or 5.46E+05 particles/mL). The digestion of milk peptides, bound preferentially over pepsin within the hard corona on the PS10 surface, was delayed up to 15 min in comparison to bulk protein digestion. Intact caseins, otherwise rapidly digested, remained bound to PS10 in the hard corona for up to 15 min. This work presents valuable insights regarding the interaction of MPs, food proteins, and pepsin, and their dynamics during gastric digestion.
PB  - Elsevier Ltd.
T2  - Environmental Pollution
T1  - Small polystyrene microplastics interfere with the breakdown of milk proteins during static in vitro simulated human gastric digestion
VL  - 335
SP  - 122282
DO  - 10.1016/j.envpol.2023.122282
ER  - 

@article{
author = "de Guzman, Maria Krishna and Stanić-Vučinić, Dragana and Gligorijević, Nikola and Wimmer, Lukas and Gasparyan, Manvel and Lujić, Tamara and Vasović, Tamara and Dailey, Lea Ann and Van Haute, Sam and Ćirković-Veličković, Tanja",
year = "2023",
abstract = "Human ingestion of microplastics (MPs) is common and inevitable due to the widespread contamination of food items, but implications on the gastric digestion of food proteins are still unknown. In this study, the interactions between pepsin and polystyrene (PS) MPs were evaluated by investigating enzyme activity and conformation in a simulated human gastric environment in the presence or absence of PS MPs. The impact on food digestion was also assessed by monitoring the kinetics of protein hydrolysis through static in vitro gastric digestion of cow's milk contaminated with PS. The binding of pepsin to PS showed that the surface chemistry of MPs dictates binding affinity. The key contributor to pepsin adsorption seems to be π−π interactions between the aromatic residues and the PS phenyl rings. During quick exposure (10 min) of pepsin to increasing concentrations (222, 2219, 22188 particles/mL) of 10 μm PS (PS10) and 100 μm PS (PS100), total enzymatic activities were not affected remarkably. However, upon prolonged exposure at 1 and 2 h, preferential binding of pepsin to the small, low zeta-potential PS caused structural changes in the protein which led to a significant reduction of its activity. Digestion of cow's milk mixed with PS10 resulted in transient accumulation of larger peptides (10–35 kDa) and reduced bioavailability of short peptides (2–9 kDa) in the gastric phase. This, however, was only observed at extremely high PS10 concentration (0.3 mg/mL or 5.46E+05 particles/mL). The digestion of milk peptides, bound preferentially over pepsin within the hard corona on the PS10 surface, was delayed up to 15 min in comparison to bulk protein digestion. Intact caseins, otherwise rapidly digested, remained bound to PS10 in the hard corona for up to 15 min. This work presents valuable insights regarding the interaction of MPs, food proteins, and pepsin, and their dynamics during gastric digestion.",
publisher = "Elsevier Ltd.",
journal = "Environmental Pollution",
title = "Small polystyrene microplastics interfere with the breakdown of milk proteins during static in vitro simulated human gastric digestion",
volume = "335",
pages = "122282",
doi = "10.1016/j.envpol.2023.122282"
}

de Guzman, M. K., Stanić-Vučinić, D., Gligorijević, N., Wimmer, L., Gasparyan, M., Lujić, T., Vasović, T., Dailey, L. A., Van Haute, S.,& Ćirković-Veličković, T.. (2023). Small polystyrene microplastics interfere with the breakdown of milk proteins during static in vitro simulated human gastric digestion. in Environmental Pollution
Elsevier Ltd.., 335, 122282.
https://doi.org/10.1016/j.envpol.2023.122282

de Guzman MK, Stanić-Vučinić D, Gligorijević N, Wimmer L, Gasparyan M, Lujić T, Vasović T, Dailey LA, Van Haute S, Ćirković-Veličković T. Small polystyrene microplastics interfere with the breakdown of milk proteins during static in vitro simulated human gastric digestion. in Environmental Pollution. 2023;335:122282.
doi:10.1016/j.envpol.2023.122282 .

de Guzman, Maria Krishna, Stanić-Vučinić, Dragana, Gligorijević, Nikola, Wimmer, Lukas, Gasparyan, Manvel, Lujić, Tamara, Vasović, Tamara, Dailey, Lea Ann, Van Haute, Sam, Ćirković-Veličković, Tanja, "Small polystyrene microplastics interfere with the breakdown of milk proteins during static in vitro simulated human gastric digestion" in Environmental Pollution, 335 (2023):122282,
https://doi.org/10.1016/j.envpol.2023.122282 . .

TY  - CONF
AU  - Radomirović, Mirjana
AU  - Gligorijević, Nikola
AU  - Minić, Simeon
AU  - Nikolić, Milan
AU  - Stanić-Vučinić, Dragana
AU  - Ćirković Veličković, Tanja
PY  - 2022
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/6945
AB  - Phycocyanobilin (PCB) is an open-chain tetrapyrrole chromophore of phycocyanin (PC), chromoprotein derived from the cyanobacterium Arthrospira platensis. Our group has previously demonstrated the potential of PCB to covalently modify free cysteine residue of proteins using food protein β-lactoglobulin as a model protein. Relying on the proven ability of PCB to be attached to sulfhydryl groups of proteins, we propose a new method for covalent attachment of PCB to potentially any protein. We used Traut’s reagent (TR, 2-iminothiolane) to introduce free sulfhydryl groups in the model protein, bovine serum albumin (BSA), by modifying its lysine residues. All tested molar ratios of TR per mole of protein successfully modified BSA. A higher degree of modification by TR induced more profound alterations of BSA structure, as evidenced by near-UV and far-UV circular dichroism spectroscopy. At the same time, minor changes in BSA oligomerization and aggregation profile occurred with increasing TR molar ratio. PCB was covalently attached to introduced sulfhydryl groups at pH 9 at 20–fold molar ratio of PCB per mole of protein. An increase in the molar ratio of TR per mole of BSA leads to amplification of fluorescent signal of PCB-modified BSA, most significantly observed starting from 50-fold and higher TR ratios. Using BSA as a model protein, a 50-fold molar excess of TR seems to be the optimal choice for balancing a satisfactory signal amplification level and the negative effect on protein structure. BSA covalently modified with PCB has higher antioxidative activity than free BSA. The proposed method thus serves as a proof of concept for labeling virtually any protein with PCB as means of either functionalization through covalent attachment of bioactive PCB or obtaining fluorescent probes for application in fluorescence-based techniques.
PB  - Wiley
PB  - Federation of European Biochemical Societies
C3  - The Biochemistry Global Summit, 9th-14th July, 2022. In: FEBS Open Bio
T1  - Covalent modification of bovine serum albumin with phycocyanobilin using Traut's reagent
VL  - 12
IS  - Suppl. 1
SP  - 302
EP  - 303
UR  - https://hdl.handle.net/21.15107/rcub_cer_6945
ER  - 

@conference{
author = "Radomirović, Mirjana and Gligorijević, Nikola and Minić, Simeon and Nikolić, Milan and Stanić-Vučinić, Dragana and Ćirković Veličković, Tanja",
year = "2022",
abstract = "Phycocyanobilin (PCB) is an open-chain tetrapyrrole chromophore of phycocyanin (PC), chromoprotein derived from the cyanobacterium Arthrospira platensis. Our group has previously demonstrated the potential of PCB to covalently modify free cysteine residue of proteins using food protein β-lactoglobulin as a model protein. Relying on the proven ability of PCB to be attached to sulfhydryl groups of proteins, we propose a new method for covalent attachment of PCB to potentially any protein. We used Traut’s reagent (TR, 2-iminothiolane) to introduce free sulfhydryl groups in the model protein, bovine serum albumin (BSA), by modifying its lysine residues. All tested molar ratios of TR per mole of protein successfully modified BSA. A higher degree of modification by TR induced more profound alterations of BSA structure, as evidenced by near-UV and far-UV circular dichroism spectroscopy. At the same time, minor changes in BSA oligomerization and aggregation profile occurred with increasing TR molar ratio. PCB was covalently attached to introduced sulfhydryl groups at pH 9 at 20–fold molar ratio of PCB per mole of protein. An increase in the molar ratio of TR per mole of BSA leads to amplification of fluorescent signal of PCB-modified BSA, most significantly observed starting from 50-fold and higher TR ratios. Using BSA as a model protein, a 50-fold molar excess of TR seems to be the optimal choice for balancing a satisfactory signal amplification level and the negative effect on protein structure. BSA covalently modified with PCB has higher antioxidative activity than free BSA. The proposed method thus serves as a proof of concept for labeling virtually any protein with PCB as means of either functionalization through covalent attachment of bioactive PCB or obtaining fluorescent probes for application in fluorescence-based techniques.",
publisher = "Wiley, Federation of European Biochemical Societies",
journal = "The Biochemistry Global Summit, 9th-14th July, 2022. In: FEBS Open Bio",
title = "Covalent modification of bovine serum albumin with phycocyanobilin using Traut's reagent",
volume = "12",
number = "Suppl. 1",
pages = "302-303",
url = "https://hdl.handle.net/21.15107/rcub_cer_6945"
}

Radomirović, M., Gligorijević, N., Minić, S., Nikolić, M., Stanić-Vučinić, D.,& Ćirković Veličković, T.. (2022). Covalent modification of bovine serum albumin with phycocyanobilin using Traut's reagent. in The Biochemistry Global Summit, 9th-14th July, 2022. In: FEBS Open Bio
Wiley., 12(Suppl. 1), 302-303.
https://hdl.handle.net/21.15107/rcub_cer_6945

Radomirović M, Gligorijević N, Minić S, Nikolić M, Stanić-Vučinić D, Ćirković Veličković T. Covalent modification of bovine serum albumin with phycocyanobilin using Traut's reagent. in The Biochemistry Global Summit, 9th-14th July, 2022. In: FEBS Open Bio. 2022;12(Suppl. 1):302-303.
https://hdl.handle.net/21.15107/rcub_cer_6945 .

Radomirović, Mirjana, Gligorijević, Nikola, Minić, Simeon, Nikolić, Milan, Stanić-Vučinić, Dragana, Ćirković Veličković, Tanja, "Covalent modification of bovine serum albumin with phycocyanobilin using Traut's reagent" in The Biochemistry Global Summit, 9th-14th July, 2022. In: FEBS Open Bio, 12, no. Suppl. 1 (2022):302-303,
https://hdl.handle.net/21.15107/rcub_cer_6945 .

TY  - CONF
AU  - Radomirović, Mirjana
AU  - Čolaković, Maša
AU  - Pismestrović, Marina
AU  - Gligorijević, Nikola
AU  - Stanić-Vučinić, Dragana
AU  - Rajković, Andreja
AU  - Ćirković Veličković, Tanja
PY  - 2022
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/6823
AB  - Tropomyosin (TPM) is considered as a major allergen among different shellfish species.
Therefore, the development of methods for quantifying TPM in food products iscrucial for
allergic persons. Several extraction buffers were tested for their efficiency in recovering
proteins from fresh frozen and cooked razor mud shrimp during 2 and 24 hours of
extraction. The protein content was quantified using the Bradford protein assay. SDSPAGE
was used for protein profiling of soluble extracts. Sandwich ELISA was developed
and used to quantify TPM content. None of the extraction buffers showed a significant
difference in total protein content between 2 and 24 hours of extraction. Significantly
fewer proteins were extracted from cooked shrimp compared to the raw shrimp. ELISA
quantification showed that phosphate-buffered saline (PBS) containing 1 M NaCl (PBSN)
and carbonate buffer, pH 10, extracted approximately 6 times higher amount of
tropomyosin in comparison to PBS, highlighting the importance of choosing the
appropriate extraction buffer for the precise quantification of TPM. Traditionally used
extraction buffer PBS could significantly underestimate shrimp TPM content.
AB  - Tropomiozin (TPM) se smatra glavnim alergenom morskih plodova. Razvoj metoda za kvantifikaciju TPM-a u prehrambenim proizvodima je ključan za alergične osobe. Nekoliko pufera za ekstrakciju je upoređeno u pogledu efikasnosti ekstrakcije protein iz sveže zamrznutih i kuvanih gambora, tokom 2 i 24 časa. Sadržaj proteina u solubilnim ekstraktima je određen Bredfordovom metodom. Proteinski profil ekstrakata je analiziran SDS-PAGE metodom. Za kvantifikaciju TPM-a je razvijena sendvič ELISA metoda. Nijedan puffer nije pokazao značajnu razliku u količini ekstrahovanih protein nakon 2 i 24 časa ekstrakcije. U poređenju sa sirovim, značajno manje proteina je ekstrahovano iz kuvanih gambora. Kvantifikacija TPM-a ELISA metodom je pokazala da se fosfatom puferisanim fiziološkim rastvorom (PBS) koji sadrži 1 M NaCl (PBSN) i karbonatnim 
puferom, pH 10, ekstrahuje oko 6 puta više TPM-a u poređenju sa PBS-om. Ovo ukazuje 
na značaj odabira adekvatnog pufera za ekstrakciju TPM-a, jer se upotrebom tradicionalno
korišćenih pufera može potceniti koncentracija TPM-a u gamborima.
PB  - Belgrade : Serbian Chemical Society
C3  - 58th Meeting of  the Serbian Chemical Society - Book of abstracts, Proceedings,  June 9-10, 2022 Belgrade / 58. Savetovanje Srpskog hemijskog društva, Kratki izvodi radova, Knjiga radova, 9. i 10. jun 2022. godine, Beograd
T1  - Optimization of extraction conditions of tropomyosin from razor mud  shrimp and its quantification by developed ELISA
SP  - 64
EP  - 64
UR  - https://hdl.handle.net/21.15107/rcub_cer_6823
ER  - 

@conference{
author = "Radomirović, Mirjana and Čolaković, Maša and Pismestrović, Marina and Gligorijević, Nikola and Stanić-Vučinić, Dragana and Rajković, Andreja and Ćirković Veličković, Tanja",
year = "2022",
abstract = "Tropomyosin (TPM) is considered as a major allergen among different shellfish species.
Therefore, the development of methods for quantifying TPM in food products iscrucial for
allergic persons. Several extraction buffers were tested for their efficiency in recovering
proteins from fresh frozen and cooked razor mud shrimp during 2 and 24 hours of
extraction. The protein content was quantified using the Bradford protein assay. SDSPAGE
was used for protein profiling of soluble extracts. Sandwich ELISA was developed
and used to quantify TPM content. None of the extraction buffers showed a significant
difference in total protein content between 2 and 24 hours of extraction. Significantly
fewer proteins were extracted from cooked shrimp compared to the raw shrimp. ELISA
quantification showed that phosphate-buffered saline (PBS) containing 1 M NaCl (PBSN)
and carbonate buffer, pH 10, extracted approximately 6 times higher amount of
tropomyosin in comparison to PBS, highlighting the importance of choosing the
appropriate extraction buffer for the precise quantification of TPM. Traditionally used
extraction buffer PBS could significantly underestimate shrimp TPM content., Tropomiozin (TPM) se smatra glavnim alergenom morskih plodova. Razvoj metoda za kvantifikaciju TPM-a u prehrambenim proizvodima je ključan za alergične osobe. Nekoliko pufera za ekstrakciju je upoređeno u pogledu efikasnosti ekstrakcije protein iz sveže zamrznutih i kuvanih gambora, tokom 2 i 24 časa. Sadržaj proteina u solubilnim ekstraktima je određen Bredfordovom metodom. Proteinski profil ekstrakata je analiziran SDS-PAGE metodom. Za kvantifikaciju TPM-a je razvijena sendvič ELISA metoda. Nijedan puffer nije pokazao značajnu razliku u količini ekstrahovanih protein nakon 2 i 24 časa ekstrakcije. U poređenju sa sirovim, značajno manje proteina je ekstrahovano iz kuvanih gambora. Kvantifikacija TPM-a ELISA metodom je pokazala da se fosfatom puferisanim fiziološkim rastvorom (PBS) koji sadrži 1 M NaCl (PBSN) i karbonatnim 
puferom, pH 10, ekstrahuje oko 6 puta više TPM-a u poređenju sa PBS-om. Ovo ukazuje 
na značaj odabira adekvatnog pufera za ekstrakciju TPM-a, jer se upotrebom tradicionalno
korišćenih pufera može potceniti koncentracija TPM-a u gamborima.",
publisher = "Belgrade : Serbian Chemical Society",
journal = "58th Meeting of  the Serbian Chemical Society - Book of abstracts, Proceedings,  June 9-10, 2022 Belgrade / 58. Savetovanje Srpskog hemijskog društva, Kratki izvodi radova, Knjiga radova, 9. i 10. jun 2022. godine, Beograd",
title = "Optimization of extraction conditions of tropomyosin from razor mud  shrimp and its quantification by developed ELISA",
pages = "64-64",
url = "https://hdl.handle.net/21.15107/rcub_cer_6823"
}

Radomirović, M., Čolaković, M., Pismestrović, M., Gligorijević, N., Stanić-Vučinić, D., Rajković, A.,& Ćirković Veličković, T.. (2022). Optimization of extraction conditions of tropomyosin from razor mud  shrimp and its quantification by developed ELISA. in 58th Meeting of  the Serbian Chemical Society - Book of abstracts, Proceedings,  June 9-10, 2022 Belgrade / 58. Savetovanje Srpskog hemijskog društva, Kratki izvodi radova, Knjiga radova, 9. i 10. jun 2022. godine, Beograd
Belgrade : Serbian Chemical Society., 64-64.
https://hdl.handle.net/21.15107/rcub_cer_6823

Radomirović M, Čolaković M, Pismestrović M, Gligorijević N, Stanić-Vučinić D, Rajković A, Ćirković Veličković T. Optimization of extraction conditions of tropomyosin from razor mud  shrimp and its quantification by developed ELISA. in 58th Meeting of  the Serbian Chemical Society - Book of abstracts, Proceedings,  June 9-10, 2022 Belgrade / 58. Savetovanje Srpskog hemijskog društva, Kratki izvodi radova, Knjiga radova, 9. i 10. jun 2022. godine, Beograd. 2022;:64-64.
https://hdl.handle.net/21.15107/rcub_cer_6823 .

Radomirović, Mirjana, Čolaković, Maša, Pismestrović, Marina, Gligorijević, Nikola, Stanić-Vučinić, Dragana, Rajković, Andreja, Ćirković Veličković, Tanja, "Optimization of extraction conditions of tropomyosin from razor mud  shrimp and its quantification by developed ELISA" in 58th Meeting of  the Serbian Chemical Society - Book of abstracts, Proceedings,  June 9-10, 2022 Belgrade / 58. Savetovanje Srpskog hemijskog društva, Kratki izvodi radova, Knjiga radova, 9. i 10. jun 2022. godine, Beograd (2022):64-64,
https://hdl.handle.net/21.15107/rcub_cer_6823 .

TY  - JOUR
AU  - Radosavljević, Jelena
AU  - Stanić-Vučinić, Dragana
AU  - Stojadinović, Marija M.
AU  - Radomirović, Mirjana Ž.
AU  - Simović, Ana
AU  - Radibratović, Milica
AU  - Ćirković-Veličković, Tanja
PY  - 2022
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/5555
AB  - Background: The world’s production of whey is estimated to be more than 200 milliontons per year. Although whey is an important source of proteins with high nutritional value andbiotechnological importance, it is still considered as a by-product of the dairy industry with loweconomic value due to low industrial exploitation. There are several challenges in the separation ofwhey proteins: low concentration, the complexity of the material and similar properties (pI, molecularmass) of some proteins.Methods: A narrative review of all the relevant papers on the present methodologies based on ionexchange and adsorption principles for isolation of whey proteins, known to the authors, was conductedResults: Traditional ion exchange techniques are widely used for the separation and purification ofthe bovine whey proteins. These methodologies, based on the anion or cation chromatographicprocedures, as well as the combination of aforementioned techniques are still preferential methodsfor the isolation of the whey proteins on the laboratory scale. However, more recent research on ionexchange membranes for this purpose has been introduced, with promising potential to be appliedon the pilot industrial scale. Newly developed methodologies based either on the ion exchangeseparation (for example: simulated moving bed chromatography, expanded bed adsorption, magneticion exchangers, etc.) or adsorption (for example: adsorption on hydroxyapatite or activatedcarbon, or molecular imprinting) are promising approaches for scaling up of the whey proteins’purification processes.Conclusion: Many procedures based on ion exchange are successfully implemented for the separationand purification of whey proteins, providing protein preparations of moderate-to-high yieldand satisfactory purity. However, the authors anticipate further development of adsorption-basedmethodologies for the separation of whey proteins by targeting the differences in proteins’ structuresrather than targeting the differences in molecular masses and pI. The complex composite multilayeredmatrices, including also inorganic components, are promising materials for simultaneousexploiting of the differences in the masses, pI and structures of whey proteins for the separation.
PB  - Bentham Science
T2  - Current Analytical Chemistry
T1  - Application of Ion Exchange and Adsorption Techniques for Separation of Whey Proteins from Bovine Milk
VL  - 18
IS  - 3
SP  - 341
EP  - 359
DO  - 10.2174/1573411017666210108092338
ER  - 

@article{
author = "Radosavljević, Jelena and Stanić-Vučinić, Dragana and Stojadinović, Marija M. and Radomirović, Mirjana Ž. and Simović, Ana and Radibratović, Milica and Ćirković-Veličković, Tanja",
year = "2022",
abstract = "Background: The world’s production of whey is estimated to be more than 200 milliontons per year. Although whey is an important source of proteins with high nutritional value andbiotechnological importance, it is still considered as a by-product of the dairy industry with loweconomic value due to low industrial exploitation. There are several challenges in the separation ofwhey proteins: low concentration, the complexity of the material and similar properties (pI, molecularmass) of some proteins.Methods: A narrative review of all the relevant papers on the present methodologies based on ionexchange and adsorption principles for isolation of whey proteins, known to the authors, was conductedResults: Traditional ion exchange techniques are widely used for the separation and purification ofthe bovine whey proteins. These methodologies, based on the anion or cation chromatographicprocedures, as well as the combination of aforementioned techniques are still preferential methodsfor the isolation of the whey proteins on the laboratory scale. However, more recent research on ionexchange membranes for this purpose has been introduced, with promising potential to be appliedon the pilot industrial scale. Newly developed methodologies based either on the ion exchangeseparation (for example: simulated moving bed chromatography, expanded bed adsorption, magneticion exchangers, etc.) or adsorption (for example: adsorption on hydroxyapatite or activatedcarbon, or molecular imprinting) are promising approaches for scaling up of the whey proteins’purification processes.Conclusion: Many procedures based on ion exchange are successfully implemented for the separationand purification of whey proteins, providing protein preparations of moderate-to-high yieldand satisfactory purity. However, the authors anticipate further development of adsorption-basedmethodologies for the separation of whey proteins by targeting the differences in proteins’ structuresrather than targeting the differences in molecular masses and pI. The complex composite multilayeredmatrices, including also inorganic components, are promising materials for simultaneousexploiting of the differences in the masses, pI and structures of whey proteins for the separation.",
publisher = "Bentham Science",
journal = "Current Analytical Chemistry",
title = "Application of Ion Exchange and Adsorption Techniques for Separation of Whey Proteins from Bovine Milk",
volume = "18",
number = "3",
pages = "341-359",
doi = "10.2174/1573411017666210108092338"
}

Radosavljević, J., Stanić-Vučinić, D., Stojadinović, M. M., Radomirović, M. Ž., Simović, A., Radibratović, M.,& Ćirković-Veličković, T.. (2022). Application of Ion Exchange and Adsorption Techniques for Separation of Whey Proteins from Bovine Milk. in Current Analytical Chemistry
Bentham Science., 18(3), 341-359.
https://doi.org/10.2174/1573411017666210108092338

Radosavljević J, Stanić-Vučinić D, Stojadinović MM, Radomirović MŽ, Simović A, Radibratović M, Ćirković-Veličković T. Application of Ion Exchange and Adsorption Techniques for Separation of Whey Proteins from Bovine Milk. in Current Analytical Chemistry. 2022;18(3):341-359.
doi:10.2174/1573411017666210108092338 .

Radosavljević, Jelena, Stanić-Vučinić, Dragana, Stojadinović, Marija M., Radomirović, Mirjana Ž., Simović, Ana, Radibratović, Milica, Ćirković-Veličković, Tanja, "Application of Ion Exchange and Adsorption Techniques for Separation of Whey Proteins from Bovine Milk" in Current Analytical Chemistry, 18, no. 3 (2022):341-359,
https://doi.org/10.2174/1573411017666210108092338 . .

TY  - JOUR
AU  - Radosavljević, Jelena
AU  - Stanić-Vučinić, Dragana
AU  - Stojadinović, Marija M.
AU  - Radomirović, Mirjana Ž.
AU  - Simović, Ana
AU  - Radibratović, Milica
AU  - Ćirković-Veličković, Tanja
PY  - 2022
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/5556
AB  - Background: The world’s production of whey is estimated to be more than 200 milliontons per year. Although whey is an important source of proteins with high nutritional value andbiotechnological importance, it is still considered as a by-product of the dairy industry with loweconomic value due to low industrial exploitation. There are several challenges in the separation ofwhey proteins: low concentration, the complexity of the material and similar properties (pI, molecularmass) of some proteins.Methods: A narrative review of all the relevant papers on the present methodologies based on ionexchange and adsorption principles for isolation of whey proteins, known to the authors, was conductedResults: Traditional ion exchange techniques are widely used for the separation and purification ofthe bovine whey proteins. These methodologies, based on the anion or cation chromatographicprocedures, as well as the combination of aforementioned techniques are still preferential methodsfor the isolation of the whey proteins on the laboratory scale. However, more recent research on ionexchange membranes for this purpose has been introduced, with promising potential to be appliedon the pilot industrial scale. Newly developed methodologies based either on the ion exchangeseparation (for example: simulated moving bed chromatography, expanded bed adsorption, magneticion exchangers, etc.) or adsorption (for example: adsorption on hydroxyapatite or activatedcarbon, or molecular imprinting) are promising approaches for scaling up of the whey proteins’purification processes.Conclusion: Many procedures based on ion exchange are successfully implemented for the separationand purification of whey proteins, providing protein preparations of moderate-to-high yieldand satisfactory purity. However, the authors anticipate further development of adsorption-basedmethodologies for the separation of whey proteins by targeting the differences in proteins’ structuresrather than targeting the differences in molecular masses and pI. The complex composite multilayeredmatrices, including also inorganic components, are promising materials for simultaneousexploiting of the differences in the masses, pI and structures of whey proteins for the separation.
PB  - Bentham Science
T2  - Current Analytical Chemistry
T1  - Application of Ion Exchange and Adsorption Techniques for Separation of Whey Proteins from Bovine Milk
VL  - 18
IS  - 3
SP  - 341
EP  - 359
DO  - 10.2174/1573411017666210108092338
ER  - 

@article{
author = "Radosavljević, Jelena and Stanić-Vučinić, Dragana and Stojadinović, Marija M. and Radomirović, Mirjana Ž. and Simović, Ana and Radibratović, Milica and Ćirković-Veličković, Tanja",
year = "2022",
abstract = "Background: The world’s production of whey is estimated to be more than 200 milliontons per year. Although whey is an important source of proteins with high nutritional value andbiotechnological importance, it is still considered as a by-product of the dairy industry with loweconomic value due to low industrial exploitation. There are several challenges in the separation ofwhey proteins: low concentration, the complexity of the material and similar properties (pI, molecularmass) of some proteins.Methods: A narrative review of all the relevant papers on the present methodologies based on ionexchange and adsorption principles for isolation of whey proteins, known to the authors, was conductedResults: Traditional ion exchange techniques are widely used for the separation and purification ofthe bovine whey proteins. These methodologies, based on the anion or cation chromatographicprocedures, as well as the combination of aforementioned techniques are still preferential methodsfor the isolation of the whey proteins on the laboratory scale. However, more recent research on ionexchange membranes for this purpose has been introduced, with promising potential to be appliedon the pilot industrial scale. Newly developed methodologies based either on the ion exchangeseparation (for example: simulated moving bed chromatography, expanded bed adsorption, magneticion exchangers, etc.) or adsorption (for example: adsorption on hydroxyapatite or activatedcarbon, or molecular imprinting) are promising approaches for scaling up of the whey proteins’purification processes.Conclusion: Many procedures based on ion exchange are successfully implemented for the separationand purification of whey proteins, providing protein preparations of moderate-to-high yieldand satisfactory purity. However, the authors anticipate further development of adsorption-basedmethodologies for the separation of whey proteins by targeting the differences in proteins’ structuresrather than targeting the differences in molecular masses and pI. The complex composite multilayeredmatrices, including also inorganic components, are promising materials for simultaneousexploiting of the differences in the masses, pI and structures of whey proteins for the separation.",
publisher = "Bentham Science",
journal = "Current Analytical Chemistry",
title = "Application of Ion Exchange and Adsorption Techniques for Separation of Whey Proteins from Bovine Milk",
volume = "18",
number = "3",
pages = "341-359",
doi = "10.2174/1573411017666210108092338"
}

Radosavljević, J., Stanić-Vučinić, D., Stojadinović, M. M., Radomirović, M. Ž., Simović, A., Radibratović, M.,& Ćirković-Veličković, T.. (2022). Application of Ion Exchange and Adsorption Techniques for Separation of Whey Proteins from Bovine Milk. in Current Analytical Chemistry
Bentham Science., 18(3), 341-359.
https://doi.org/10.2174/1573411017666210108092338

Radosavljević J, Stanić-Vučinić D, Stojadinović MM, Radomirović MŽ, Simović A, Radibratović M, Ćirković-Veličković T. Application of Ion Exchange and Adsorption Techniques for Separation of Whey Proteins from Bovine Milk. in Current Analytical Chemistry. 2022;18(3):341-359.
doi:10.2174/1573411017666210108092338 .

Radosavljević, Jelena, Stanić-Vučinić, Dragana, Stojadinović, Marija M., Radomirović, Mirjana Ž., Simović, Ana, Radibratović, Milica, Ćirković-Veličković, Tanja, "Application of Ion Exchange and Adsorption Techniques for Separation of Whey Proteins from Bovine Milk" in Current Analytical Chemistry, 18, no. 3 (2022):341-359,
https://doi.org/10.2174/1573411017666210108092338 . .

TY  - CONF
AU  - Radomirović, Mirjana
AU  - Gligorijević, Nikola
AU  - Minić, Simeon
AU  - Nikolić, Milan
AU  - Stanić-Vučinić, Dragana
AU  - Ćirković-Veličković, Tanja
PY  - 2021
UR  - https://ec.europa.eu/research/participants/documents/downloadPublic?documentIds=080166e5deeb546e&appId=PPGMS
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/7282
AB  - Phycobiliproteins (PBP) have been employed in numerous fluorescence-based techniques owing
to highly fluorescent, covalently bound tetrapyrrole chromophores. So far, only entire PBPs have
been utilized as fluorescent probes. A new method for covalent attachment of phycocyanin’s
chromophore, phycocyanobilin (PCB), to potentially any protein, is proposed, relying on the
ability of PCB to be attached to sulfhydryl groups of proteins. Traut’s reagent (TR, 2-
iminothiolane) was exploited for introduction of sulfhydryl groups in the model protein, bovine
serum albumin (BSA), by modifying its primary amines. Introduced sulfhydryl groups were then
targeted for modification by PCB. All tested molar ratios of TR per mole of protein were
successful in modification of BSA. Near-UV and far-UV circular dichroism spectroscopy
revealed that a higher degree of modification by TR induces more profound alterations of BSA
structure, leading at the same time to minor changes in BSA oligomerization and aggregation
profile. PCB was covalently attached to introduced sulfhydryl groups at pH 9 at 20–fold ratio of
TR. An increase in the molar ratio of TR per mole of BSA leads to amplification of fluorescent
signal of PCB-modified BSA, most significantly observed starting from 50-fold and higher TR
ratios. Using BSA as a model protein, a 50-fold molar excess of TR seems to be the optimal ratio
for balancing between the effect on protein structure and the degree of labeling and thus
fluorescent signal obtained. The proposed method could be used for labeling of virtually any
protein, as means of either obtaining fluorescent probes for application in fluorescent techniques
or functionalization of, for example, food proteins through covalent attachment of bioactive
PCB.
PB  - University of Belgrade - Faculty of Chemistry
C3  - FoodEnTwin Symposium “Novel analytical approaches in food and environmental sciences”, Book of Abstracts, 16th-18th June, 2021, Belgrade, Serbia
T1  - Traut’s reagent application in fluorescent labeling of bovine serum albumin with phycocyanobilin
SP  - 37
EP  - 37
UR  - https://hdl.handle.net/21.15107/rcub_cer_7282
ER  - 

@conference{
author = "Radomirović, Mirjana and Gligorijević, Nikola and Minić, Simeon and Nikolić, Milan and Stanić-Vučinić, Dragana and Ćirković-Veličković, Tanja",
year = "2021",
abstract = "Phycobiliproteins (PBP) have been employed in numerous fluorescence-based techniques owing
to highly fluorescent, covalently bound tetrapyrrole chromophores. So far, only entire PBPs have
been utilized as fluorescent probes. A new method for covalent attachment of phycocyanin’s
chromophore, phycocyanobilin (PCB), to potentially any protein, is proposed, relying on the
ability of PCB to be attached to sulfhydryl groups of proteins. Traut’s reagent (TR, 2-
iminothiolane) was exploited for introduction of sulfhydryl groups in the model protein, bovine
serum albumin (BSA), by modifying its primary amines. Introduced sulfhydryl groups were then
targeted for modification by PCB. All tested molar ratios of TR per mole of protein were
successful in modification of BSA. Near-UV and far-UV circular dichroism spectroscopy
revealed that a higher degree of modification by TR induces more profound alterations of BSA
structure, leading at the same time to minor changes in BSA oligomerization and aggregation
profile. PCB was covalently attached to introduced sulfhydryl groups at pH 9 at 20–fold ratio of
TR. An increase in the molar ratio of TR per mole of BSA leads to amplification of fluorescent
signal of PCB-modified BSA, most significantly observed starting from 50-fold and higher TR
ratios. Using BSA as a model protein, a 50-fold molar excess of TR seems to be the optimal ratio
for balancing between the effect on protein structure and the degree of labeling and thus
fluorescent signal obtained. The proposed method could be used for labeling of virtually any
protein, as means of either obtaining fluorescent probes for application in fluorescent techniques
or functionalization of, for example, food proteins through covalent attachment of bioactive
PCB.",
publisher = "University of Belgrade - Faculty of Chemistry",
journal = "FoodEnTwin Symposium “Novel analytical approaches in food and environmental sciences”, Book of Abstracts, 16th-18th June, 2021, Belgrade, Serbia",
title = "Traut’s reagent application in fluorescent labeling of bovine serum albumin with phycocyanobilin",
pages = "37-37",
url = "https://hdl.handle.net/21.15107/rcub_cer_7282"
}

Radomirović, M., Gligorijević, N., Minić, S., Nikolić, M., Stanić-Vučinić, D.,& Ćirković-Veličković, T.. (2021). Traut’s reagent application in fluorescent labeling of bovine serum albumin with phycocyanobilin. in FoodEnTwin Symposium “Novel analytical approaches in food and environmental sciences”, Book of Abstracts, 16th-18th June, 2021, Belgrade, Serbia
University of Belgrade - Faculty of Chemistry., 37-37.
https://hdl.handle.net/21.15107/rcub_cer_7282

Radomirović M, Gligorijević N, Minić S, Nikolić M, Stanić-Vučinić D, Ćirković-Veličković T. Traut’s reagent application in fluorescent labeling of bovine serum albumin with phycocyanobilin. in FoodEnTwin Symposium “Novel analytical approaches in food and environmental sciences”, Book of Abstracts, 16th-18th June, 2021, Belgrade, Serbia. 2021;:37-37.
https://hdl.handle.net/21.15107/rcub_cer_7282 .

Radomirović, Mirjana, Gligorijević, Nikola, Minić, Simeon, Nikolić, Milan, Stanić-Vučinić, Dragana, Ćirković-Veličković, Tanja, "Traut’s reagent application in fluorescent labeling of bovine serum albumin with phycocyanobilin" in FoodEnTwin Symposium “Novel analytical approaches in food and environmental sciences”, Book of Abstracts, 16th-18th June, 2021, Belgrade, Serbia (2021):37-37,
https://hdl.handle.net/21.15107/rcub_cer_7282 .

TY  - CONF
AU  - Radomirović, Mirjana
AU  - Gligorijević, Nikola
AU  - Minić, Simeon
AU  - Nikolić, Milan
AU  - Stanić-Vučinić, Dragana
AU  - Ćirković Veličković, Tanja
PY  - 2021
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/7290
AB  - Phycobiliproteins (PBP) are extensively used as fluorescent probes due to highly fluorescent,
covalently bound, tetrapyrrole chromophores. A new method for covalent attachment of
phycocyanin’s chromophore, phycocyanobilin (PCB), is proposed. We exploited Traut’s reagent
(TR) to introduce sulfhydryl groups in the bovine serum albumin (BSA), by modifying its lysine
residues. TR successfully modified BSA under all tested molar ratios of reagent per mole of BSA.
The higher degree of modification by TR induces more profound alterations of BSA structure. PCB
was covalently attached to introduced sulfhydryl groups at pH 9 at 20–fold ratio. An increase in the
molar ratio of TR per mole of BSA leads to amplification of fluorescent signal of PCB-modified
BSA. Using BSA as a model protein, a 50-fold molar excess of TR seems to be the optimal choice
for balancing between a satisfactory level of signal amplification and the adverse effect on protein
structure. The method could be used for labeling virtually any protein.
PB  - Belgrade : Serbian Chemical Society
C3  - Kratki izvodi radova, Knjiga radova 57. Savetovanje Srpskog hemijskog društva, 18. i 19. juni 2021, Kragujevac / Book of abstracts, Proceedings - 57th Meeting of the Serbian Chemical Society, June 18-19, 2021, Kragujevac, Serbia
T1  - Fluorescent labeling of bovine serum albumin with phycocyanobilin using Traut’s reagent
SP  - 71
EP  - 71
UR  - https://hdl.handle.net/21.15107/rcub_cer_7290
ER  - 

@conference{
author = "Radomirović, Mirjana and Gligorijević, Nikola and Minić, Simeon and Nikolić, Milan and Stanić-Vučinić, Dragana and Ćirković Veličković, Tanja",
year = "2021",
abstract = "Phycobiliproteins (PBP) are extensively used as fluorescent probes due to highly fluorescent,
covalently bound, tetrapyrrole chromophores. A new method for covalent attachment of
phycocyanin’s chromophore, phycocyanobilin (PCB), is proposed. We exploited Traut’s reagent
(TR) to introduce sulfhydryl groups in the bovine serum albumin (BSA), by modifying its lysine
residues. TR successfully modified BSA under all tested molar ratios of reagent per mole of BSA.
The higher degree of modification by TR induces more profound alterations of BSA structure. PCB
was covalently attached to introduced sulfhydryl groups at pH 9 at 20–fold ratio. An increase in the
molar ratio of TR per mole of BSA leads to amplification of fluorescent signal of PCB-modified
BSA. Using BSA as a model protein, a 50-fold molar excess of TR seems to be the optimal choice
for balancing between a satisfactory level of signal amplification and the adverse effect on protein
structure. The method could be used for labeling virtually any protein.",
publisher = "Belgrade : Serbian Chemical Society",
journal = "Kratki izvodi radova, Knjiga radova 57. Savetovanje Srpskog hemijskog društva, 18. i 19. juni 2021, Kragujevac / Book of abstracts, Proceedings - 57th Meeting of the Serbian Chemical Society, June 18-19, 2021, Kragujevac, Serbia",
title = "Fluorescent labeling of bovine serum albumin with phycocyanobilin using Traut’s reagent",
pages = "71-71",
url = "https://hdl.handle.net/21.15107/rcub_cer_7290"
}

Radomirović, M., Gligorijević, N., Minić, S., Nikolić, M., Stanić-Vučinić, D.,& Ćirković Veličković, T.. (2021). Fluorescent labeling of bovine serum albumin with phycocyanobilin using Traut’s reagent. in Kratki izvodi radova, Knjiga radova 57. Savetovanje Srpskog hemijskog društva, 18. i 19. juni 2021, Kragujevac / Book of abstracts, Proceedings - 57th Meeting of the Serbian Chemical Society, June 18-19, 2021, Kragujevac, Serbia
Belgrade : Serbian Chemical Society., 71-71.
https://hdl.handle.net/21.15107/rcub_cer_7290

Radomirović M, Gligorijević N, Minić S, Nikolić M, Stanić-Vučinić D, Ćirković Veličković T. Fluorescent labeling of bovine serum albumin with phycocyanobilin using Traut’s reagent. in Kratki izvodi radova, Knjiga radova 57. Savetovanje Srpskog hemijskog društva, 18. i 19. juni 2021, Kragujevac / Book of abstracts, Proceedings - 57th Meeting of the Serbian Chemical Society, June 18-19, 2021, Kragujevac, Serbia. 2021;:71-71.
https://hdl.handle.net/21.15107/rcub_cer_7290 .

Radomirović, Mirjana, Gligorijević, Nikola, Minić, Simeon, Nikolić, Milan, Stanić-Vučinić, Dragana, Ćirković Veličković, Tanja, "Fluorescent labeling of bovine serum albumin with phycocyanobilin using Traut’s reagent" in Kratki izvodi radova, Knjiga radova 57. Savetovanje Srpskog hemijskog društva, 18. i 19. juni 2021, Kragujevac / Book of abstracts, Proceedings - 57th Meeting of the Serbian Chemical Society, June 18-19, 2021, Kragujevac, Serbia (2021):71-71,
https://hdl.handle.net/21.15107/rcub_cer_7290 .

TY  - CONF
AU  - Radomirović, Mirjana
AU  - Gligorijević, Nikola
AU  - Stanić-Vučinić, Dragana
AU  - Rajković, Andreja
AU  - Ćirković Veličković, Tanja
PY  - 2021
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/7280
AB  - Food allergies affect up to 10% of the general population and represent an important health problem in the field of
food safety in industrialized countries. Hence, developing reliable, specific, and sensitive methods for detecting and
quantifying allergens in food products is of high importance. Shellfish have been recognized as one of the eight
most common sources of allergens, with tropomyosin (TPM) being considered a major heat-stable allergen, having
a highly conserved amino acid sequence among different shellfish species. Allergenicity of TPM may change during
food processing, such as cooking. The objective of this study was to develop an enzyme-linked immunosorbent
assay (ELISA) for the detection and quantification of shellfish tropomyosin in food samples.
Two different extraction buffers - phosphate-buffered saline (PBS) and PBS containing 1 M sodium-chloride
(PBSN), were compared for their ability to recover proteins from pre-cooked frozen Mediterranean mussel (Mytilus
galloprovincialis) and fresh frozen razor mud shrimp (Solenocera melantho). The samples were additionally cooked
according to the manufacturer's instruction and analyzed as such. The protein content was quantified using Bradford
protein assay, and the protein components of soluble extracts were profiled using SDS-PAGE. TPM presence was
confirmed using Western blot. Sandwich ELISA was developed using a monoclonal anti-TPM antibody as a capture
antibody, while polyclonal anti-TPM antibody served as a detection antibody and was coupled to the biotinylated
secondary antibody and streptavidin-alkaline phosphatase conjugate. Tropomyosin was quantified using highly
purified natural shrimp tropomyosin as standard.
The profile of extracted proteins was changed when using PBSN instead of PBS. A higher concentration of proteins
was recovered from raw shrimp using PBSN instead of PBS. At the same time, the type of extraction buffer did not
affect protein recovery either from heated shrimp or pre-cooked/heated mussels. Significantly fewer proteins were
extracted from cooked shrimp sample compared to the raw shrimp, while cooking showed no effect on the extraction
of proteins from mussels. Cooking did not affect TPM recognition in Western blot. TPM was quantified in shrimp
samples in sandwich ELISA. However, developed ELISA could not quantify mussel's TPM, indicating that this
approach may distinguish mussels and shrimp TPM.
PB  - Sociedade Portuguesa de Química
C3  - Book of Abstracts - XXI EuroFoodChem conference, 22nd-24th November, 2021, Virtual Congress
T1  - Extraction and quantification of tropomyosin in selected samples of shellfish
SP  - 118
EP  - 118
UR  - https://hdl.handle.net/21.15107/rcub_cer_7280
ER  - 

@conference{
author = "Radomirović, Mirjana and Gligorijević, Nikola and Stanić-Vučinić, Dragana and Rajković, Andreja and Ćirković Veličković, Tanja",
year = "2021",
abstract = "Food allergies affect up to 10% of the general population and represent an important health problem in the field of
food safety in industrialized countries. Hence, developing reliable, specific, and sensitive methods for detecting and
quantifying allergens in food products is of high importance. Shellfish have been recognized as one of the eight
most common sources of allergens, with tropomyosin (TPM) being considered a major heat-stable allergen, having
a highly conserved amino acid sequence among different shellfish species. Allergenicity of TPM may change during
food processing, such as cooking. The objective of this study was to develop an enzyme-linked immunosorbent
assay (ELISA) for the detection and quantification of shellfish tropomyosin in food samples.
Two different extraction buffers - phosphate-buffered saline (PBS) and PBS containing 1 M sodium-chloride
(PBSN), were compared for their ability to recover proteins from pre-cooked frozen Mediterranean mussel (Mytilus
galloprovincialis) and fresh frozen razor mud shrimp (Solenocera melantho). The samples were additionally cooked
according to the manufacturer's instruction and analyzed as such. The protein content was quantified using Bradford
protein assay, and the protein components of soluble extracts were profiled using SDS-PAGE. TPM presence was
confirmed using Western blot. Sandwich ELISA was developed using a monoclonal anti-TPM antibody as a capture
antibody, while polyclonal anti-TPM antibody served as a detection antibody and was coupled to the biotinylated
secondary antibody and streptavidin-alkaline phosphatase conjugate. Tropomyosin was quantified using highly
purified natural shrimp tropomyosin as standard.
The profile of extracted proteins was changed when using PBSN instead of PBS. A higher concentration of proteins
was recovered from raw shrimp using PBSN instead of PBS. At the same time, the type of extraction buffer did not
affect protein recovery either from heated shrimp or pre-cooked/heated mussels. Significantly fewer proteins were
extracted from cooked shrimp sample compared to the raw shrimp, while cooking showed no effect on the extraction
of proteins from mussels. Cooking did not affect TPM recognition in Western blot. TPM was quantified in shrimp
samples in sandwich ELISA. However, developed ELISA could not quantify mussel's TPM, indicating that this
approach may distinguish mussels and shrimp TPM.",
publisher = "Sociedade Portuguesa de Química",
journal = "Book of Abstracts - XXI EuroFoodChem conference, 22nd-24th November, 2021, Virtual Congress",
title = "Extraction and quantification of tropomyosin in selected samples of shellfish",
pages = "118-118",
url = "https://hdl.handle.net/21.15107/rcub_cer_7280"
}

Radomirović, M., Gligorijević, N., Stanić-Vučinić, D., Rajković, A.,& Ćirković Veličković, T.. (2021). Extraction and quantification of tropomyosin in selected samples of shellfish. in Book of Abstracts - XXI EuroFoodChem conference, 22nd-24th November, 2021, Virtual Congress
Sociedade Portuguesa de Química., 118-118.
https://hdl.handle.net/21.15107/rcub_cer_7280

Radomirović M, Gligorijević N, Stanić-Vučinić D, Rajković A, Ćirković Veličković T. Extraction and quantification of tropomyosin in selected samples of shellfish. in Book of Abstracts - XXI EuroFoodChem conference, 22nd-24th November, 2021, Virtual Congress. 2021;:118-118.
https://hdl.handle.net/21.15107/rcub_cer_7280 .

Radomirović, Mirjana, Gligorijević, Nikola, Stanić-Vučinić, Dragana, Rajković, Andreja, Ćirković Veličković, Tanja, "Extraction and quantification of tropomyosin in selected samples of shellfish" in Book of Abstracts - XXI EuroFoodChem conference, 22nd-24th November, 2021, Virtual Congress (2021):118-118,
https://hdl.handle.net/21.15107/rcub_cer_7280 .

TY  - JOUR
AU  - Radibratović, Milica
AU  - Al-Hanish, Ayah
AU  - Minić, Simeon
AU  - Radomirović, Mirjana
AU  - Milčić, Miloš
AU  - Stanić-Vučinić, Dragana
AU  - Ćirković Veličković, Tanja
PY  - 2019
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/3212
AB  - α-Lactalbumin (ALA) is a Ca2+-binding protein which constitutes up to 20% of whey protein. At acidic pH, and in the apo-state at elevated temperatures, ALA is the classic 'molten globule' (MG). This study examined epigallocatechin-3-gallate (EGCG) binding to ALA in its apo form (apoALA) and stabilizing effect on protein structure thereof.

EGCG binds to apoALA in both native and MG state. The complex of EGCG and ALA is more stable to thermal denaturation. The docking analysis and molecular dynamic simulation (MDS) showed that Ca2+ removal decreased conformational stability of ALA, because of the local destabilization of Ca2+-binding region. EGCG binding to apoALA increases its stability by reverting of conformation and stability of Ca2+-binding region. Therefore, EGCG-induced thermal stability of apoALA is based on increased apoALA conformational rigidity. This study implies that during gastric digestion of tea with milk EGCG would remain bound to ALA, albeit in the Ca2+-free form
PB  - Elsevier
T2  - Food Chemistry
T1  - Stabilization of apo α-lactalbumin by binding of epigallocatechin-3-gallate: Experimental and molecular dynamics study
VL  - 278
SP  - 388
EP  - 395
DO  - 10.1016/j.foodchem.2018.11.038
ER  - 

@article{
author = "Radibratović, Milica and Al-Hanish, Ayah and Minić, Simeon and Radomirović, Mirjana and Milčić, Miloš and Stanić-Vučinić, Dragana and Ćirković Veličković, Tanja",
year = "2019",
abstract = "α-Lactalbumin (ALA) is a Ca2+-binding protein which constitutes up to 20% of whey protein. At acidic pH, and in the apo-state at elevated temperatures, ALA is the classic 'molten globule' (MG). This study examined epigallocatechin-3-gallate (EGCG) binding to ALA in its apo form (apoALA) and stabilizing effect on protein structure thereof.

EGCG binds to apoALA in both native and MG state. The complex of EGCG and ALA is more stable to thermal denaturation. The docking analysis and molecular dynamic simulation (MDS) showed that Ca2+ removal decreased conformational stability of ALA, because of the local destabilization of Ca2+-binding region. EGCG binding to apoALA increases its stability by reverting of conformation and stability of Ca2+-binding region. Therefore, EGCG-induced thermal stability of apoALA is based on increased apoALA conformational rigidity. This study implies that during gastric digestion of tea with milk EGCG would remain bound to ALA, albeit in the Ca2+-free form",
publisher = "Elsevier",
journal = "Food Chemistry",
title = "Stabilization of apo α-lactalbumin by binding of epigallocatechin-3-gallate: Experimental and molecular dynamics study",
volume = "278",
pages = "388-395",
doi = "10.1016/j.foodchem.2018.11.038"
}

Radibratović, M., Al-Hanish, A., Minić, S., Radomirović, M., Milčić, M., Stanić-Vučinić, D.,& Ćirković Veličković, T.. (2019). Stabilization of apo α-lactalbumin by binding of epigallocatechin-3-gallate: Experimental and molecular dynamics study. in Food Chemistry
Elsevier., 278, 388-395.
https://doi.org/10.1016/j.foodchem.2018.11.038

Radibratović M, Al-Hanish A, Minić S, Radomirović M, Milčić M, Stanić-Vučinić D, Ćirković Veličković T. Stabilization of apo α-lactalbumin by binding of epigallocatechin-3-gallate: Experimental and molecular dynamics study. in Food Chemistry. 2019;278:388-395.
doi:10.1016/j.foodchem.2018.11.038 .

Radibratović, Milica, Al-Hanish, Ayah, Minić, Simeon, Radomirović, Mirjana, Milčić, Miloš, Stanić-Vučinić, Dragana, Ćirković Veličković, Tanja, "Stabilization of apo α-lactalbumin by binding of epigallocatechin-3-gallate: Experimental and molecular dynamics study" in Food Chemistry, 278 (2019):388-395,
https://doi.org/10.1016/j.foodchem.2018.11.038 . .

TY  - JOUR
AU  - Minić, Simeon
AU  - Radomirović, Mirjana
AU  - Savkovic, Nina
AU  - Radibratović, Milica
AU  - Mihailović, Jelena
AU  - Vasović, Tamara
AU  - Nikolić, Milan
AU  - Milčić, Miloš
AU  - Stanić-Vučinić, Dragana
AU  - Ćirković Veličković, Tanja
PY  - 2018
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/2420
AB  - In this study, we investigated structural aspects of covalent binding of food derived blue pigment phycocyanobilin (PCB) to bovine beta-lactoglobulin (BLG), major whey protein, by spectroscopic, electrophoretic, mass spectrometry and computational methods. At physiological pH (7.2), we found that covalent pigment binding via free cysteine residue is slow (k(a)=0.065 min(-1)), of moderate affinity (K-a=4x10(4) M-1), and stereo-selective. Binding also occurs at a broad pH range and under simulated gastrointestinal conditions. Adduct formation rises with pH, and in concentrated urea (k(a)=0.101 min(-1)). The BLG-PCB adduct has slightly altered secondary and tertiary protein structure, and bound PCB has higher fluorescence and more stretched conformation than free chromophore. Combination of steered molecular dynamic for disulfide exchange, non-covalent and covalent docking, favours Cys119 residue in protein calyx as target for covalent BLG-PCB adduct formation. Our results suggest that this adduct can serve as delivery system of bioactive PCB.
PB  - Elsevier Sci Ltd, Oxford
T2  - Food Chemistry
T1  - Covalent binding of food-derived blue pigment phycocyanobilin to bovine beta-lactoglobulin under physiological conditions
VL  - 269
SP  - 43
EP  - 52
DO  - 10.1016/j.foodchem.2018.06.138
ER  - 

@article{
author = "Minić, Simeon and Radomirović, Mirjana and Savkovic, Nina and Radibratović, Milica and Mihailović, Jelena and Vasović, Tamara and Nikolić, Milan and Milčić, Miloš and Stanić-Vučinić, Dragana and Ćirković Veličković, Tanja",
year = "2018",
abstract = "In this study, we investigated structural aspects of covalent binding of food derived blue pigment phycocyanobilin (PCB) to bovine beta-lactoglobulin (BLG), major whey protein, by spectroscopic, electrophoretic, mass spectrometry and computational methods. At physiological pH (7.2), we found that covalent pigment binding via free cysteine residue is slow (k(a)=0.065 min(-1)), of moderate affinity (K-a=4x10(4) M-1), and stereo-selective. Binding also occurs at a broad pH range and under simulated gastrointestinal conditions. Adduct formation rises with pH, and in concentrated urea (k(a)=0.101 min(-1)). The BLG-PCB adduct has slightly altered secondary and tertiary protein structure, and bound PCB has higher fluorescence and more stretched conformation than free chromophore. Combination of steered molecular dynamic for disulfide exchange, non-covalent and covalent docking, favours Cys119 residue in protein calyx as target for covalent BLG-PCB adduct formation. Our results suggest that this adduct can serve as delivery system of bioactive PCB.",
publisher = "Elsevier Sci Ltd, Oxford",
journal = "Food Chemistry",
title = "Covalent binding of food-derived blue pigment phycocyanobilin to bovine beta-lactoglobulin under physiological conditions",
volume = "269",
pages = "43-52",
doi = "10.1016/j.foodchem.2018.06.138"
}

Minić, S., Radomirović, M., Savkovic, N., Radibratović, M., Mihailović, J., Vasović, T., Nikolić, M., Milčić, M., Stanić-Vučinić, D.,& Ćirković Veličković, T.. (2018). Covalent binding of food-derived blue pigment phycocyanobilin to bovine beta-lactoglobulin under physiological conditions. in Food Chemistry
Elsevier Sci Ltd, Oxford., 269, 43-52.
https://doi.org/10.1016/j.foodchem.2018.06.138

Minić S, Radomirović M, Savkovic N, Radibratović M, Mihailović J, Vasović T, Nikolić M, Milčić M, Stanić-Vučinić D, Ćirković Veličković T. Covalent binding of food-derived blue pigment phycocyanobilin to bovine beta-lactoglobulin under physiological conditions. in Food Chemistry. 2018;269:43-52.
doi:10.1016/j.foodchem.2018.06.138 .

Minić, Simeon, Radomirović, Mirjana, Savkovic, Nina, Radibratović, Milica, Mihailović, Jelena, Vasović, Tamara, Nikolić, Milan, Milčić, Miloš, Stanić-Vučinić, Dragana, Ćirković Veličković, Tanja, "Covalent binding of food-derived blue pigment phycocyanobilin to bovine beta-lactoglobulin under physiological conditions" in Food Chemistry, 269 (2018):43-52,
https://doi.org/10.1016/j.foodchem.2018.06.138 . .

TY  - JOUR
AU  - Minić, Simeon
AU  - Stanić-Vučinić, Dragana
AU  - Radomirović, Mirjana
AU  - Radibratović, Milica
AU  - Milčić, Miloš
AU  - Nikolić, Milan
AU  - Ćirković Veličković, Tanja
PY  - 2018
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/2478
AB  - Phycocyanobilin (PCB) is a blue tetrapyrrole chromophore of C-phycocyanin, the main protein of the microalga Spirulina, with numerous proven health-related benefits. We examined binding of PCB to bovine serum albumin (BSA) and how it affects protein and ligand stability. Protein fluorescence quenching and microscale thermophoresis demonstrated high-affinity binding (K-a = 2 x 10(6) M-1). Spectroscopic titration with molecular docking analysis revealed two binding sites on BSA, at the inter-domain cleft and at subdomain IB, while CD spectroscopy indicated stereo-selective binding of the P conformer of the pigment to the protein. The PCB protein complex showed increased thermal stability. Although complex formation partly masked the antioxidant properties of PCB and BSA, a mutually protective effect against free radical-induced oxidation was found. BSA could be suitable for delivery of PCB as a food colorant or bioactive component. Our results also highlight subtle differences between PCB binding to bovine vs. human serum albumin.
PB  - Elsevier Sci Ltd, Oxford
T2  - Food Chemistry
T1  - Characterization and effects of binding of food-derived bioactive phycocyanobilin to bovine serum albumin
VL  - 239
SP  - 1090
EP  - 1099
DO  - 10.1016/j.foodchem.2017.07.066
ER  - 

@article{
author = "Minić, Simeon and Stanić-Vučinić, Dragana and Radomirović, Mirjana and Radibratović, Milica and Milčić, Miloš and Nikolić, Milan and Ćirković Veličković, Tanja",
year = "2018",
abstract = "Phycocyanobilin (PCB) is a blue tetrapyrrole chromophore of C-phycocyanin, the main protein of the microalga Spirulina, with numerous proven health-related benefits. We examined binding of PCB to bovine serum albumin (BSA) and how it affects protein and ligand stability. Protein fluorescence quenching and microscale thermophoresis demonstrated high-affinity binding (K-a = 2 x 10(6) M-1). Spectroscopic titration with molecular docking analysis revealed two binding sites on BSA, at the inter-domain cleft and at subdomain IB, while CD spectroscopy indicated stereo-selective binding of the P conformer of the pigment to the protein. The PCB protein complex showed increased thermal stability. Although complex formation partly masked the antioxidant properties of PCB and BSA, a mutually protective effect against free radical-induced oxidation was found. BSA could be suitable for delivery of PCB as a food colorant or bioactive component. Our results also highlight subtle differences between PCB binding to bovine vs. human serum albumin.",
publisher = "Elsevier Sci Ltd, Oxford",
journal = "Food Chemistry",
title = "Characterization and effects of binding of food-derived bioactive phycocyanobilin to bovine serum albumin",
volume = "239",
pages = "1090-1099",
doi = "10.1016/j.foodchem.2017.07.066"
}

Minić, S., Stanić-Vučinić, D., Radomirović, M., Radibratović, M., Milčić, M., Nikolić, M.,& Ćirković Veličković, T.. (2018). Characterization and effects of binding of food-derived bioactive phycocyanobilin to bovine serum albumin. in Food Chemistry
Elsevier Sci Ltd, Oxford., 239, 1090-1099.
https://doi.org/10.1016/j.foodchem.2017.07.066

Minić S, Stanić-Vučinić D, Radomirović M, Radibratović M, Milčić M, Nikolić M, Ćirković Veličković T. Characterization and effects of binding of food-derived bioactive phycocyanobilin to bovine serum albumin. in Food Chemistry. 2018;239:1090-1099.
doi:10.1016/j.foodchem.2017.07.066 .

Minić, Simeon, Stanić-Vučinić, Dragana, Radomirović, Mirjana, Radibratović, Milica, Milčić, Miloš, Nikolić, Milan, Ćirković Veličković, Tanja, "Characterization and effects of binding of food-derived bioactive phycocyanobilin to bovine serum albumin" in Food Chemistry, 239 (2018):1090-1099,
https://doi.org/10.1016/j.foodchem.2017.07.066 . .

TY  - JOUR
AU  - Minić, Simeon
AU  - Stanić-Vučinić, Dragana
AU  - Radomirović, Mirjana
AU  - Radibratović, Milica
AU  - Milčić, Miloš
AU  - Nikolić, Milan
AU  - Ćirković Veličković, Tanja
PY  - 2018
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/2984
AB  - Phycocyanobilin (PCB) is a blue tetrapyrrole chromophore of C-phycocyanin, the main protein of the microalga Spirulina, with numerous proven health-related benefits. We examined binding of PCB to bovine serum albumin (BSA) and how it affects protein and ligand stability. Protein fluorescence quenching and microscale thermophoresis demonstrated high-affinity binding (K-a = 2 x 10(6) M-1). Spectroscopic titration with molecular docking analysis revealed two binding sites on BSA, at the inter-domain cleft and at subdomain IB, while CD spectroscopy indicated stereo-selective binding of the P conformer of the pigment to the protein. The PCB protein complex showed increased thermal stability. Although complex formation partly masked the antioxidant properties of PCB and BSA, a mutually protective effect against free radical-induced oxidation was found. BSA could be suitable for delivery of PCB as a food colorant or bioactive component. Our results also highlight subtle differences between PCB binding to bovine vs. human serum albumin.
PB  - Elsevier Sci Ltd, Oxford
T2  - Food Chemistry
T1  - Characterization and effects of binding of food-derived bioactive phycocyanobilin to bovine serum albumin
VL  - 239
SP  - 1090
EP  - 1099
DO  - 10.1016/j.foodchem.2017.07.066
ER  - 

@article{
author = "Minić, Simeon and Stanić-Vučinić, Dragana and Radomirović, Mirjana and Radibratović, Milica and Milčić, Miloš and Nikolić, Milan and Ćirković Veličković, Tanja",
year = "2018",
abstract = "Phycocyanobilin (PCB) is a blue tetrapyrrole chromophore of C-phycocyanin, the main protein of the microalga Spirulina, with numerous proven health-related benefits. We examined binding of PCB to bovine serum albumin (BSA) and how it affects protein and ligand stability. Protein fluorescence quenching and microscale thermophoresis demonstrated high-affinity binding (K-a = 2 x 10(6) M-1). Spectroscopic titration with molecular docking analysis revealed two binding sites on BSA, at the inter-domain cleft and at subdomain IB, while CD spectroscopy indicated stereo-selective binding of the P conformer of the pigment to the protein. The PCB protein complex showed increased thermal stability. Although complex formation partly masked the antioxidant properties of PCB and BSA, a mutually protective effect against free radical-induced oxidation was found. BSA could be suitable for delivery of PCB as a food colorant or bioactive component. Our results also highlight subtle differences between PCB binding to bovine vs. human serum albumin.",
publisher = "Elsevier Sci Ltd, Oxford",
journal = "Food Chemistry",
title = "Characterization and effects of binding of food-derived bioactive phycocyanobilin to bovine serum albumin",
volume = "239",
pages = "1090-1099",
doi = "10.1016/j.foodchem.2017.07.066"
}

Minić, S., Stanić-Vučinić, D., Radomirović, M., Radibratović, M., Milčić, M., Nikolić, M.,& Ćirković Veličković, T.. (2018). Characterization and effects of binding of food-derived bioactive phycocyanobilin to bovine serum albumin. in Food Chemistry
Elsevier Sci Ltd, Oxford., 239, 1090-1099.
https://doi.org/10.1016/j.foodchem.2017.07.066

Minić S, Stanić-Vučinić D, Radomirović M, Radibratović M, Milčić M, Nikolić M, Ćirković Veličković T. Characterization and effects of binding of food-derived bioactive phycocyanobilin to bovine serum albumin. in Food Chemistry. 2018;239:1090-1099.
doi:10.1016/j.foodchem.2017.07.066 .

Minić, Simeon, Stanić-Vučinić, Dragana, Radomirović, Mirjana, Radibratović, Milica, Milčić, Miloš, Nikolić, Milan, Ćirković Veličković, Tanja, "Characterization and effects of binding of food-derived bioactive phycocyanobilin to bovine serum albumin" in Food Chemistry, 239 (2018):1090-1099,
https://doi.org/10.1016/j.foodchem.2017.07.066 . .

TY  - JOUR
AU  - Minić, Simeon
AU  - Radomirović, Mirjana
AU  - Savkovic, Nina
AU  - Radibratović, Milica
AU  - Mihailović, Jelena
AU  - Vasović, Tamara
AU  - Nikolić, Milan
AU  - Milčić, Miloš
AU  - Stanić-Vučinić, Dragana
AU  - Ćirković Veličković, Tanja
PY  - 2018
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/3022
AB  - In this study, we investigated structural aspects of covalent binding of food derived blue pigment phycocyanobilin (PCB) to bovine beta-lactoglobulin (BLG), major whey protein, by spectroscopic, electrophoretic, mass spectrometry and computational methods. At physiological pH (7.2), we found that covalent pigment binding via free cysteine residue is slow (k(a)=0.065 min(-1)), of moderate affinity (K-a=4x10(4) M-1), and stereo-selective. Binding also occurs at a broad pH range and under simulated gastrointestinal conditions. Adduct formation rises with pH, and in concentrated urea (k(a)=0.101 min(-1)). The BLG-PCB adduct has slightly altered secondary and tertiary protein structure, and bound PCB has higher fluorescence and more stretched conformation than free chromophore. Combination of steered molecular dynamic for disulfide exchange, non-covalent and covalent docking, favours Cys119 residue in protein calyx as target for covalent BLG-PCB adduct formation. Our results suggest that this adduct can serve as delivery system of bioactive PCB.
PB  - Elsevier Sci Ltd, Oxford
T2  - Food Chemistry
T1  - Covalent binding of food-derived blue pigment phycocyanobilin to bovine beta-lactoglobulin under physiological conditions
VL  - 269
SP  - 43
EP  - 52
DO  - 10.1016/j.foodchem.2018.06.138
ER  - 

@article{
author = "Minić, Simeon and Radomirović, Mirjana and Savkovic, Nina and Radibratović, Milica and Mihailović, Jelena and Vasović, Tamara and Nikolić, Milan and Milčić, Miloš and Stanić-Vučinić, Dragana and Ćirković Veličković, Tanja",
year = "2018",
abstract = "In this study, we investigated structural aspects of covalent binding of food derived blue pigment phycocyanobilin (PCB) to bovine beta-lactoglobulin (BLG), major whey protein, by spectroscopic, electrophoretic, mass spectrometry and computational methods. At physiological pH (7.2), we found that covalent pigment binding via free cysteine residue is slow (k(a)=0.065 min(-1)), of moderate affinity (K-a=4x10(4) M-1), and stereo-selective. Binding also occurs at a broad pH range and under simulated gastrointestinal conditions. Adduct formation rises with pH, and in concentrated urea (k(a)=0.101 min(-1)). The BLG-PCB adduct has slightly altered secondary and tertiary protein structure, and bound PCB has higher fluorescence and more stretched conformation than free chromophore. Combination of steered molecular dynamic for disulfide exchange, non-covalent and covalent docking, favours Cys119 residue in protein calyx as target for covalent BLG-PCB adduct formation. Our results suggest that this adduct can serve as delivery system of bioactive PCB.",
publisher = "Elsevier Sci Ltd, Oxford",
journal = "Food Chemistry",
title = "Covalent binding of food-derived blue pigment phycocyanobilin to bovine beta-lactoglobulin under physiological conditions",
volume = "269",
pages = "43-52",
doi = "10.1016/j.foodchem.2018.06.138"
}

Minić, S., Radomirović, M., Savkovic, N., Radibratović, M., Mihailović, J., Vasović, T., Nikolić, M., Milčić, M., Stanić-Vučinić, D.,& Ćirković Veličković, T.. (2018). Covalent binding of food-derived blue pigment phycocyanobilin to bovine beta-lactoglobulin under physiological conditions. in Food Chemistry
Elsevier Sci Ltd, Oxford., 269, 43-52.
https://doi.org/10.1016/j.foodchem.2018.06.138

Minić S, Radomirović M, Savkovic N, Radibratović M, Mihailović J, Vasović T, Nikolić M, Milčić M, Stanić-Vučinić D, Ćirković Veličković T. Covalent binding of food-derived blue pigment phycocyanobilin to bovine beta-lactoglobulin under physiological conditions. in Food Chemistry. 2018;269:43-52.
doi:10.1016/j.foodchem.2018.06.138 .

Minić, Simeon, Radomirović, Mirjana, Savkovic, Nina, Radibratović, Milica, Mihailović, Jelena, Vasović, Tamara, Nikolić, Milan, Milčić, Miloš, Stanić-Vučinić, Dragana, Ćirković Veličković, Tanja, "Covalent binding of food-derived blue pigment phycocyanobilin to bovine beta-lactoglobulin under physiological conditions" in Food Chemistry, 269 (2018):43-52,
https://doi.org/10.1016/j.foodchem.2018.06.138 . .

TY  - CONF
AU  - Radibratović, Milica
AU  - Al-Hanish, Ayah
AU  - Minić, Simeon
AU  - Radomirović, Mirjana Ž.
AU  - Milčić, Miloš
AU  - Stanić-Vučinić, Dragana
AU  - Ćirković-Veličković, Tanja
PY  - 2018
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/7462
AB  - α-Lactalbumin (ALA) is a small, acidic, Ca2+-binding protein that might constitute up to 20% of whey protein. ALA has several partially folded intermediate states. It is very attractive for studies of the properties and structure of intermediate molten globule-like states since at acidic pH and in the apo-state at elevated temperatures ALA is the classic ‘molten globule’.The aim of our study was to examine if epi-gallo-catechin-3-gallate (EGCG) binds to ALA in its Ca-depleted (apo-ALA) and less ordered states. Moreover we examined if the polyphenol binding can stabilize protein structure. The experimental investigation of EGCG binding and protein’s stability were supported by molecular dynamics data.Green tea catechin binds to apo-ALA in neutral and acidic conditions. Binding affinity determined by intrinsic fluorescence quenching is of a similar magnitude as to the holo form of the protein. The complex of EGCG and ALA at neutral conditions is more stable to thermal denaturation then protein alone. The stabilizing effect is more pronounced for apo-ALA than for the holo-ALA.The docking analysis and molecular dynamic simulation (MDS) showed that EGCG binds to apoALA at the same site as to holoALA, in the hydrophobic pocket at the entrance of cleft between α and β subdomains, remaining bound during MDS. Ca2+ removal results in decreased conformational stability of ALA, where local destabilization of Ca2+-binding region propagates to cleft, with its slight opening. EGCG binding to apo-ALA increases stability of ALA by reverting of conformation and stability of disturbed Ca2+-binding region, which is transmitted to cleft, resulting in rejoining of α and β subdomains and slight cleft closing by the same mechanism. EGCG binding could retard transition of apo-ALA to less ordered states under conditions favoring its formation.Therefore, non-covalent binding of EGCG can stabilize structural fold of calcium-depleted ALA.
PB  - Univerzitet u Beogradu
C3  - UNIFOOD Conference, Belgrade, Serbia, 5th–6th October, 2018. In: UNIFOOD Conference - Programme and Book of Abstracts
T1  - Stabilization of apo-alpha-lactalbumin by binding of epigallocatechin-3-gallate: experimental and molecular dynamics study
SP  - 268
EP  - 268
UR  - https://hdl.handle.net/21.15107/rcub_cer_7462
ER  - 

@conference{
author = "Radibratović, Milica and Al-Hanish, Ayah and Minić, Simeon and Radomirović, Mirjana Ž. and Milčić, Miloš and Stanić-Vučinić, Dragana and Ćirković-Veličković, Tanja",
year = "2018",
abstract = "α-Lactalbumin (ALA) is a small, acidic, Ca2+-binding protein that might constitute up to 20% of whey protein. ALA has several partially folded intermediate states. It is very attractive for studies of the properties and structure of intermediate molten globule-like states since at acidic pH and in the apo-state at elevated temperatures ALA is the classic ‘molten globule’.The aim of our study was to examine if epi-gallo-catechin-3-gallate (EGCG) binds to ALA in its Ca-depleted (apo-ALA) and less ordered states. Moreover we examined if the polyphenol binding can stabilize protein structure. The experimental investigation of EGCG binding and protein’s stability were supported by molecular dynamics data.Green tea catechin binds to apo-ALA in neutral and acidic conditions. Binding affinity determined by intrinsic fluorescence quenching is of a similar magnitude as to the holo form of the protein. The complex of EGCG and ALA at neutral conditions is more stable to thermal denaturation then protein alone. The stabilizing effect is more pronounced for apo-ALA than for the holo-ALA.The docking analysis and molecular dynamic simulation (MDS) showed that EGCG binds to apoALA at the same site as to holoALA, in the hydrophobic pocket at the entrance of cleft between α and β subdomains, remaining bound during MDS. Ca2+ removal results in decreased conformational stability of ALA, where local destabilization of Ca2+-binding region propagates to cleft, with its slight opening. EGCG binding to apo-ALA increases stability of ALA by reverting of conformation and stability of disturbed Ca2+-binding region, which is transmitted to cleft, resulting in rejoining of α and β subdomains and slight cleft closing by the same mechanism. EGCG binding could retard transition of apo-ALA to less ordered states under conditions favoring its formation.Therefore, non-covalent binding of EGCG can stabilize structural fold of calcium-depleted ALA.",
publisher = "Univerzitet u Beogradu",
journal = "UNIFOOD Conference, Belgrade, Serbia, 5th–6th October, 2018. In: UNIFOOD Conference - Programme and Book of Abstracts",
title = "Stabilization of apo-alpha-lactalbumin by binding of epigallocatechin-3-gallate: experimental and molecular dynamics study",
pages = "268-268",
url = "https://hdl.handle.net/21.15107/rcub_cer_7462"
}

Radibratović, M., Al-Hanish, A., Minić, S., Radomirović, M. Ž., Milčić, M., Stanić-Vučinić, D.,& Ćirković-Veličković, T.. (2018). Stabilization of apo-alpha-lactalbumin by binding of epigallocatechin-3-gallate: experimental and molecular dynamics study. in UNIFOOD Conference, Belgrade, Serbia, 5th–6th October, 2018. In: UNIFOOD Conference - Programme and Book of Abstracts
Univerzitet u Beogradu., 268-268.
https://hdl.handle.net/21.15107/rcub_cer_7462

Radibratović M, Al-Hanish A, Minić S, Radomirović MŽ, Milčić M, Stanić-Vučinić D, Ćirković-Veličković T. Stabilization of apo-alpha-lactalbumin by binding of epigallocatechin-3-gallate: experimental and molecular dynamics study. in UNIFOOD Conference, Belgrade, Serbia, 5th–6th October, 2018. In: UNIFOOD Conference - Programme and Book of Abstracts. 2018;:268-268.
https://hdl.handle.net/21.15107/rcub_cer_7462 .

Radibratović, Milica, Al-Hanish, Ayah, Minić, Simeon, Radomirović, Mirjana Ž., Milčić, Miloš, Stanić-Vučinić, Dragana, Ćirković-Veličković, Tanja, "Stabilization of apo-alpha-lactalbumin by binding of epigallocatechin-3-gallate: experimental and molecular dynamics study" in UNIFOOD Conference, Belgrade, Serbia, 5th–6th October, 2018. In: UNIFOOD Conference - Programme and Book of Abstracts (2018):268-268,
https://hdl.handle.net/21.15107/rcub_cer_7462 .

TY  - JOUR
AU  - Prodić, I.
AU  - Stanić-Vučinić, Dragana
AU  - Apostolovic, D.
AU  - Mihailović, Jelena
AU  - Radibratović, Milica
AU  - Radosavljevic, J.
AU  - Burazer, L.
AU  - Milčić, Miloš
AU  - Smiljanic, K.
AU  - van, Hage M.
AU  - Ćirković Veličković, Tanja
PY  - 2018
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/2299
AB  - BackgroundMost food allergens sensitizing via the gastrointestinal tract are stable proteins that are resistant to pepsin digestion, in particular major peanut allergens, Ara h 2 and Ara h 6. Survival of their large fragments is essential for sensitizing capacity. However, the immunoreactive proteins/peptides to which the immune system of the gastrointestinal tract is exposed during digestion of peanut proteins are unknown. Particularly, the IgE reactivity of short digestion-resistant peptides (SDRPs;  LT 10kDa) released by gastric digestion under standardized and physiologically relevant invitro conditions has not been investigated. ObjectiveThe aim of this study was to investigate and identify digestion products of major peanut allergens and in particular to examine IgE reactivity of SDRPs released by pepsin digestion of whole peanut grains. MethodsTwo-dimensional gel-based proteomics and shotgun peptidomics, immunoblotting with allergen-specific antibodies from peanut-sensitized patients, enzyme-linked immunosorbent inhibition assay and ImmunoCAP tests, including far ultraviolet-circular dichroism spectroscopy were used to identify and characterize peanut digesta. ResultsAra h 2 and Ara h 6 remained mostly intact, and SDRPs from Ara h 2 were more potent in inhibiting IgE binding than Ara h 1 and Ara 3. Ara h 1 and Ara h 3 exhibited sequential digestion into a series of digestion-resistant peptides with preserved allergenic capacity. A high number of identified SDRPs from Ara h 1, Ara h 2 and Ara h 3 were part of short continuous epitope sequences and possessed substantial allergenic potential. Conclusion and Clinical RelevancePeanut grain digestion by oral and gastric phase enzymes generates mixture of products, where the major peanut allergens remain intact and their digested peptides have preserved allergenic capacity highlighting their important roles in allergic reactions to peanut.
PB  - Wiley, Hoboken
T2  - Clinical and Experimental Allergy
T1  - Influence of peanut matrix on stability of allergens in gastric-simulated digesta: 2S albumins are main contributors to the IgE reactivity of short digestion-resistant peptides
VL  - 48
IS  - 6
SP  - 731
EP  - 740
DO  - 10.1111/cea.13113
ER  - 

@article{
author = "Prodić, I. and Stanić-Vučinić, Dragana and Apostolovic, D. and Mihailović, Jelena and Radibratović, Milica and Radosavljevic, J. and Burazer, L. and Milčić, Miloš and Smiljanic, K. and van, Hage M. and Ćirković Veličković, Tanja",
year = "2018",
abstract = "BackgroundMost food allergens sensitizing via the gastrointestinal tract are stable proteins that are resistant to pepsin digestion, in particular major peanut allergens, Ara h 2 and Ara h 6. Survival of their large fragments is essential for sensitizing capacity. However, the immunoreactive proteins/peptides to which the immune system of the gastrointestinal tract is exposed during digestion of peanut proteins are unknown. Particularly, the IgE reactivity of short digestion-resistant peptides (SDRPs;  LT 10kDa) released by gastric digestion under standardized and physiologically relevant invitro conditions has not been investigated. ObjectiveThe aim of this study was to investigate and identify digestion products of major peanut allergens and in particular to examine IgE reactivity of SDRPs released by pepsin digestion of whole peanut grains. MethodsTwo-dimensional gel-based proteomics and shotgun peptidomics, immunoblotting with allergen-specific antibodies from peanut-sensitized patients, enzyme-linked immunosorbent inhibition assay and ImmunoCAP tests, including far ultraviolet-circular dichroism spectroscopy were used to identify and characterize peanut digesta. ResultsAra h 2 and Ara h 6 remained mostly intact, and SDRPs from Ara h 2 were more potent in inhibiting IgE binding than Ara h 1 and Ara 3. Ara h 1 and Ara h 3 exhibited sequential digestion into a series of digestion-resistant peptides with preserved allergenic capacity. A high number of identified SDRPs from Ara h 1, Ara h 2 and Ara h 3 were part of short continuous epitope sequences and possessed substantial allergenic potential. Conclusion and Clinical RelevancePeanut grain digestion by oral and gastric phase enzymes generates mixture of products, where the major peanut allergens remain intact and their digested peptides have preserved allergenic capacity highlighting their important roles in allergic reactions to peanut.",
publisher = "Wiley, Hoboken",
journal = "Clinical and Experimental Allergy",
title = "Influence of peanut matrix on stability of allergens in gastric-simulated digesta: 2S albumins are main contributors to the IgE reactivity of short digestion-resistant peptides",
volume = "48",
number = "6",
pages = "731-740",
doi = "10.1111/cea.13113"
}

Prodić, I., Stanić-Vučinić, D., Apostolovic, D., Mihailović, J., Radibratović, M., Radosavljevic, J., Burazer, L., Milčić, M., Smiljanic, K., van, H. M.,& Ćirković Veličković, T.. (2018). Influence of peanut matrix on stability of allergens in gastric-simulated digesta: 2S albumins are main contributors to the IgE reactivity of short digestion-resistant peptides. in Clinical and Experimental Allergy
Wiley, Hoboken., 48(6), 731-740.
https://doi.org/10.1111/cea.13113

Prodić I, Stanić-Vučinić D, Apostolovic D, Mihailović J, Radibratović M, Radosavljevic J, Burazer L, Milčić M, Smiljanic K, van HM, Ćirković Veličković T. Influence of peanut matrix on stability of allergens in gastric-simulated digesta: 2S albumins are main contributors to the IgE reactivity of short digestion-resistant peptides. in Clinical and Experimental Allergy. 2018;48(6):731-740.
doi:10.1111/cea.13113 .

Prodić, I., Stanić-Vučinić, Dragana, Apostolovic, D., Mihailović, Jelena, Radibratović, Milica, Radosavljevic, J., Burazer, L., Milčić, Miloš, Smiljanic, K., van, Hage M., Ćirković Veličković, Tanja, "Influence of peanut matrix on stability of allergens in gastric-simulated digesta: 2S albumins are main contributors to the IgE reactivity of short digestion-resistant peptides" in Clinical and Experimental Allergy, 48, no. 6 (2018):731-740,
https://doi.org/10.1111/cea.13113 . .

TY  - JOUR
AU  - Prodić, I.
AU  - Stanić-Vučinić, Dragana
AU  - Apostolovic, D.
AU  - Mihailović, Jelena
AU  - Radibratović, Milica
AU  - Radosavljevic, J.
AU  - Burazer, L.
AU  - Milčić, Miloš
AU  - Smiljanic, K.
AU  - van, Hage M.
AU  - Ćirković Veličković, Tanja
PY  - 2018
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/3340
AB  - BackgroundMost food allergens sensitizing via the gastrointestinal tract are stable proteins that are resistant to pepsin digestion, in particular major peanut allergens, Ara h 2 and Ara h 6. Survival of their large fragments is essential for sensitizing capacity. However, the immunoreactive proteins/peptides to which the immune system of the gastrointestinal tract is exposed during digestion of peanut proteins are unknown. Particularly, the IgE reactivity of short digestion-resistant peptides (SDRPs;  LT 10kDa) released by gastric digestion under standardized and physiologically relevant invitro conditions has not been investigated. ObjectiveThe aim of this study was to investigate and identify digestion products of major peanut allergens and in particular to examine IgE reactivity of SDRPs released by pepsin digestion of whole peanut grains. MethodsTwo-dimensional gel-based proteomics and shotgun peptidomics, immunoblotting with allergen-specific antibodies from peanut-sensitized patients, enzyme-linked immunosorbent inhibition assay and ImmunoCAP tests, including far ultraviolet-circular dichroism spectroscopy were used to identify and characterize peanut digesta. ResultsAra h 2 and Ara h 6 remained mostly intact, and SDRPs from Ara h 2 were more potent in inhibiting IgE binding than Ara h 1 and Ara 3. Ara h 1 and Ara h 3 exhibited sequential digestion into a series of digestion-resistant peptides with preserved allergenic capacity. A high number of identified SDRPs from Ara h 1, Ara h 2 and Ara h 3 were part of short continuous epitope sequences and possessed substantial allergenic potential. Conclusion and Clinical RelevancePeanut grain digestion by oral and gastric phase enzymes generates mixture of products, where the major peanut allergens remain intact and their digested peptides have preserved allergenic capacity highlighting their important roles in allergic reactions to peanut.
PB  - Wiley, Hoboken
T2  - Clinical and Experimental Allergy
T1  - Influence of peanut matrix on stability of allergens in gastric-simulated digesta: 2S albumins are main contributors to the IgE reactivity of short digestion-resistant peptides
VL  - 48
IS  - 6
SP  - 731
EP  - 740
DO  - 10.1111/cea.13113
ER  - 

@article{
author = "Prodić, I. and Stanić-Vučinić, Dragana and Apostolovic, D. and Mihailović, Jelena and Radibratović, Milica and Radosavljevic, J. and Burazer, L. and Milčić, Miloš and Smiljanic, K. and van, Hage M. and Ćirković Veličković, Tanja",
year = "2018",
abstract = "BackgroundMost food allergens sensitizing via the gastrointestinal tract are stable proteins that are resistant to pepsin digestion, in particular major peanut allergens, Ara h 2 and Ara h 6. Survival of their large fragments is essential for sensitizing capacity. However, the immunoreactive proteins/peptides to which the immune system of the gastrointestinal tract is exposed during digestion of peanut proteins are unknown. Particularly, the IgE reactivity of short digestion-resistant peptides (SDRPs;  LT 10kDa) released by gastric digestion under standardized and physiologically relevant invitro conditions has not been investigated. ObjectiveThe aim of this study was to investigate and identify digestion products of major peanut allergens and in particular to examine IgE reactivity of SDRPs released by pepsin digestion of whole peanut grains. MethodsTwo-dimensional gel-based proteomics and shotgun peptidomics, immunoblotting with allergen-specific antibodies from peanut-sensitized patients, enzyme-linked immunosorbent inhibition assay and ImmunoCAP tests, including far ultraviolet-circular dichroism spectroscopy were used to identify and characterize peanut digesta. ResultsAra h 2 and Ara h 6 remained mostly intact, and SDRPs from Ara h 2 were more potent in inhibiting IgE binding than Ara h 1 and Ara 3. Ara h 1 and Ara h 3 exhibited sequential digestion into a series of digestion-resistant peptides with preserved allergenic capacity. A high number of identified SDRPs from Ara h 1, Ara h 2 and Ara h 3 were part of short continuous epitope sequences and possessed substantial allergenic potential. Conclusion and Clinical RelevancePeanut grain digestion by oral and gastric phase enzymes generates mixture of products, where the major peanut allergens remain intact and their digested peptides have preserved allergenic capacity highlighting their important roles in allergic reactions to peanut.",
publisher = "Wiley, Hoboken",
journal = "Clinical and Experimental Allergy",
title = "Influence of peanut matrix on stability of allergens in gastric-simulated digesta: 2S albumins are main contributors to the IgE reactivity of short digestion-resistant peptides",
volume = "48",
number = "6",
pages = "731-740",
doi = "10.1111/cea.13113"
}

Prodić, I., Stanić-Vučinić, D., Apostolovic, D., Mihailović, J., Radibratović, M., Radosavljevic, J., Burazer, L., Milčić, M., Smiljanic, K., van, H. M.,& Ćirković Veličković, T.. (2018). Influence of peanut matrix on stability of allergens in gastric-simulated digesta: 2S albumins are main contributors to the IgE reactivity of short digestion-resistant peptides. in Clinical and Experimental Allergy
Wiley, Hoboken., 48(6), 731-740.
https://doi.org/10.1111/cea.13113

Prodić I, Stanić-Vučinić D, Apostolovic D, Mihailović J, Radibratović M, Radosavljevic J, Burazer L, Milčić M, Smiljanic K, van HM, Ćirković Veličković T. Influence of peanut matrix on stability of allergens in gastric-simulated digesta: 2S albumins are main contributors to the IgE reactivity of short digestion-resistant peptides. in Clinical and Experimental Allergy. 2018;48(6):731-740.
doi:10.1111/cea.13113 .

Prodić, I., Stanić-Vučinić, Dragana, Apostolovic, D., Mihailović, Jelena, Radibratović, Milica, Radosavljevic, J., Burazer, L., Milčić, Miloš, Smiljanic, K., van, Hage M., Ćirković Veličković, Tanja, "Influence of peanut matrix on stability of allergens in gastric-simulated digesta: 2S albumins are main contributors to the IgE reactivity of short digestion-resistant peptides" in Clinical and Experimental Allergy, 48, no. 6 (2018):731-740,
https://doi.org/10.1111/cea.13113 . .

TY  - CONF
AU  - Minic, S.
AU  - Radomirović, Mirjana
AU  - Radibratović, Milica
AU  - Milčić, Miloš
AU  - Stanić-Vučinić, Dragana
AU  - Nikolić, Milan
AU  - Ćirković Veličković, Tanja
PY  - 2017
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/2167
UR  - https://doi.org/10.1111/febs.14174
PB  - Wiley, Hoboken
C3  - FEBS Journal
T1  - Characterization and effects of binding of food-derived bioactive phycocyanobilin to bovine serum albumin
VL  - 284
IS  - S1
SP  - 189
EP  - 190
UR  - https://hdl.handle.net/21.15107/rcub_cer_2167
ER  - 

@conference{
author = "Minic, S. and Radomirović, Mirjana and Radibratović, Milica and Milčić, Miloš and Stanić-Vučinić, Dragana and Nikolić, Milan and Ćirković Veličković, Tanja",
year = "2017",
publisher = "Wiley, Hoboken",
journal = "FEBS Journal",
title = "Characterization and effects of binding of food-derived bioactive phycocyanobilin to bovine serum albumin",
volume = "284",
number = "S1",
pages = "189-190",
url = "https://hdl.handle.net/21.15107/rcub_cer_2167"
}

Minic, S., Radomirović, M., Radibratović, M., Milčić, M., Stanić-Vučinić, D., Nikolić, M.,& Ćirković Veličković, T.. (2017). Characterization and effects of binding of food-derived bioactive phycocyanobilin to bovine serum albumin. in FEBS Journal
Wiley, Hoboken., 284(S1), 189-190.
https://hdl.handle.net/21.15107/rcub_cer_2167

Minic S, Radomirović M, Radibratović M, Milčić M, Stanić-Vučinić D, Nikolić M, Ćirković Veličković T. Characterization and effects of binding of food-derived bioactive phycocyanobilin to bovine serum albumin. in FEBS Journal. 2017;284(S1):189-190.
https://hdl.handle.net/21.15107/rcub_cer_2167 .

Minic, S., Radomirović, Mirjana, Radibratović, Milica, Milčić, Miloš, Stanić-Vučinić, Dragana, Nikolić, Milan, Ćirković Veličković, Tanja, "Characterization and effects of binding of food-derived bioactive phycocyanobilin to bovine serum albumin" in FEBS Journal, 284, no. S1 (2017):189-190,
https://hdl.handle.net/21.15107/rcub_cer_2167 .

TY  - JOUR
AU  - Radibratović, Milica
AU  - Minić, Simeon
AU  - Stanić-Vučinić, Dragana
AU  - Nikolić, Milan
AU  - Milčić, Miloš
AU  - Ćirković Veličković, Tanja
PY  - 2016
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/1891
AB  - Phycocyanobilin (PCB) binds with high affinity (2.2 x 10 6 M-1 at 25 degrees C) to human serum albumin (HSA) at sites located in IB and IIA subdomains. The aim of this study was to examine effects of PCB binding on protein conformation and stability. Using 300 ns molecular dynamics (MD) simulations, UV-VIS spectrophotometry, CD, FT-IR, spectrofluorimetry, thermal denaturation and susceptibility to trypsin digestion, we studied the effects of PCB binding on the stability and rigidity of HSA, as well as the conformational changes in PCB itself upon binding to the protein. MD simulation results demonstrated that HSA with PCB bound at any of the two sites showed greater rigidity and lower overall and individual domain flexibility compared to free HSA. Experimental data demonstrated an increase in the a-helical content of the protein and thermal and proteolytic stability upon ligand binding. PCB bound to HSA undergoes a conformational change to a more elongated conformation in the binding pockets of HSA. PCB binding to HSA stabilizes the structure of this flexible transport protein, making it more thermostable and resistant to proteolysis. The results from this work explain at molecular level, conformational changes and stabilization of HSA structure upon ligand binding. The resultant increased thermal and proteolytic stability of HSA may provide greater longevity to HSA in plasma.
PB  - Public Library Science, San Francisco
T2  - Plos One
T1  - Stabilization of Human Serum Albumin by the Binding of Phycocyanobilin, a Bioactive Chromophore of Blue-Green Alga Spirulina: Molecular Dynamics and Experimental Study
VL  - 11
IS  - 12
DO  - 10.1371/journal.pone.0167973
ER  - 

@article{
author = "Radibratović, Milica and Minić, Simeon and Stanić-Vučinić, Dragana and Nikolić, Milan and Milčić, Miloš and Ćirković Veličković, Tanja",
year = "2016",
abstract = "Phycocyanobilin (PCB) binds with high affinity (2.2 x 10 6 M-1 at 25 degrees C) to human serum albumin (HSA) at sites located in IB and IIA subdomains. The aim of this study was to examine effects of PCB binding on protein conformation and stability. Using 300 ns molecular dynamics (MD) simulations, UV-VIS spectrophotometry, CD, FT-IR, spectrofluorimetry, thermal denaturation and susceptibility to trypsin digestion, we studied the effects of PCB binding on the stability and rigidity of HSA, as well as the conformational changes in PCB itself upon binding to the protein. MD simulation results demonstrated that HSA with PCB bound at any of the two sites showed greater rigidity and lower overall and individual domain flexibility compared to free HSA. Experimental data demonstrated an increase in the a-helical content of the protein and thermal and proteolytic stability upon ligand binding. PCB bound to HSA undergoes a conformational change to a more elongated conformation in the binding pockets of HSA. PCB binding to HSA stabilizes the structure of this flexible transport protein, making it more thermostable and resistant to proteolysis. The results from this work explain at molecular level, conformational changes and stabilization of HSA structure upon ligand binding. The resultant increased thermal and proteolytic stability of HSA may provide greater longevity to HSA in plasma.",
publisher = "Public Library Science, San Francisco",
journal = "Plos One",
title = "Stabilization of Human Serum Albumin by the Binding of Phycocyanobilin, a Bioactive Chromophore of Blue-Green Alga Spirulina: Molecular Dynamics and Experimental Study",
volume = "11",
number = "12",
doi = "10.1371/journal.pone.0167973"
}

Radibratović, M., Minić, S., Stanić-Vučinić, D., Nikolić, M., Milčić, M.,& Ćirković Veličković, T.. (2016). Stabilization of Human Serum Albumin by the Binding of Phycocyanobilin, a Bioactive Chromophore of Blue-Green Alga Spirulina: Molecular Dynamics and Experimental Study. in Plos One
Public Library Science, San Francisco., 11(12).
https://doi.org/10.1371/journal.pone.0167973

Radibratović M, Minić S, Stanić-Vučinić D, Nikolić M, Milčić M, Ćirković Veličković T. Stabilization of Human Serum Albumin by the Binding of Phycocyanobilin, a Bioactive Chromophore of Blue-Green Alga Spirulina: Molecular Dynamics and Experimental Study. in Plos One. 2016;11(12).
doi:10.1371/journal.pone.0167973 .

Radibratović, Milica, Minić, Simeon, Stanić-Vučinić, Dragana, Nikolić, Milan, Milčić, Miloš, Ćirković Veličković, Tanja, "Stabilization of Human Serum Albumin by the Binding of Phycocyanobilin, a Bioactive Chromophore of Blue-Green Alga Spirulina: Molecular Dynamics and Experimental Study" in Plos One, 11, no. 12 (2016),
https://doi.org/10.1371/journal.pone.0167973 . .