TY  - JOUR
AU  - Balaž, Ana Marija
AU  - Stevanović, Jelena
AU  - Ostafe, Raluca
AU  - Blažić, Marija
AU  - Ilić Đurđić, Karla
AU  - Fischer, Rainer
AU  - Prodanović, Radivoje
PY  - 2020
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/3128
AB  - Cellobiose dehydrogenase (CDH, EC 1.1.99.18) from white rot fungi Phanerochaete chrysosporium can be used for constructing biosensors and biofuel cells, for bleaching cotton in textile industry, and recently, the enzyme has found an important application in biomedicine as an antimicrobial and antibiofilm agent. Stability and activity of the wild-type (wt) CDH and mutants at methionine residues in the presence of hydrogen peroxide were investigated. Saturation mutagenesis libraries were made at the only methionine in heme domain M65 and two methionines M685 and M738 in the flavin domain that were closest to the active site. After screening the libraries, three mutants with increased activity and stability in the presence of peroxide were found, M65F with 70% of residual activity after 6 h of incubation in 0.3 M hydrogen peroxide, M738S with 80% of residual activity and M685Y with over 90% of residual activity compared to wild-type CDH that retained 40% of original activity. Combined mutants showed no activity. The most stable mutant M685Y with 5.8 times increased half-life in the presence of peroxide showed also 2.5 times increased kcat for lactose compared to wtCDH and could be good candidate for applications in biofuel cells and biocatalysis for lactobionic acid production.
PB  - Springer International Publishing
T2  - Molecular Diversity
T1  - Semi‑rational design of cellobiose dehydrogenase for increased stability in the presence of peroxide
VL  - 24
SP  - 593
EP  - 601
DO  - 10.1007/s11030-019-09965-0
ER  - 

@article{
author = "Balaž, Ana Marija and Stevanović, Jelena and Ostafe, Raluca and Blažić, Marija and Ilić Đurđić, Karla and Fischer, Rainer and Prodanović, Radivoje",
year = "2020",
abstract = "Cellobiose dehydrogenase (CDH, EC 1.1.99.18) from white rot fungi Phanerochaete chrysosporium can be used for constructing biosensors and biofuel cells, for bleaching cotton in textile industry, and recently, the enzyme has found an important application in biomedicine as an antimicrobial and antibiofilm agent. Stability and activity of the wild-type (wt) CDH and mutants at methionine residues in the presence of hydrogen peroxide were investigated. Saturation mutagenesis libraries were made at the only methionine in heme domain M65 and two methionines M685 and M738 in the flavin domain that were closest to the active site. After screening the libraries, three mutants with increased activity and stability in the presence of peroxide were found, M65F with 70% of residual activity after 6 h of incubation in 0.3 M hydrogen peroxide, M738S with 80% of residual activity and M685Y with over 90% of residual activity compared to wild-type CDH that retained 40% of original activity. Combined mutants showed no activity. The most stable mutant M685Y with 5.8 times increased half-life in the presence of peroxide showed also 2.5 times increased kcat for lactose compared to wtCDH and could be good candidate for applications in biofuel cells and biocatalysis for lactobionic acid production.",
publisher = "Springer International Publishing",
journal = "Molecular Diversity",
title = "Semi‑rational design of cellobiose dehydrogenase for increased stability in the presence of peroxide",
volume = "24",
pages = "593-601",
doi = "10.1007/s11030-019-09965-0"
}

Balaž, A. M., Stevanović, J., Ostafe, R., Blažić, M., Ilić Đurđić, K., Fischer, R.,& Prodanović, R.. (2020). Semi‑rational design of cellobiose dehydrogenase for increased stability in the presence of peroxide. in Molecular Diversity
Springer International Publishing., 24, 593-601.
https://doi.org/10.1007/s11030-019-09965-0

Balaž AM, Stevanović J, Ostafe R, Blažić M, Ilić Đurđić K, Fischer R, Prodanović R. Semi‑rational design of cellobiose dehydrogenase for increased stability in the presence of peroxide. in Molecular Diversity. 2020;24:593-601.
doi:10.1007/s11030-019-09965-0 .

Balaž, Ana Marija, Stevanović, Jelena, Ostafe, Raluca, Blažić, Marija, Ilić Đurđić, Karla, Fischer, Rainer, Prodanović, Radivoje, "Semi‑rational design of cellobiose dehydrogenase for increased stability in the presence of peroxide" in Molecular Diversity, 24 (2020):593-601,
https://doi.org/10.1007/s11030-019-09965-0 . .

TY  - JOUR
AU  - Ilić Đurđić, Karla
AU  - Ece, Selin
AU  - Ostafe, Raluca
AU  - Vogel, Simon
AU  - Balaž, Ana Marija
AU  - Schillberg, Stefan
AU  - Fischer, Rainer
AU  - Prodanović, Radivoje
PY  - 2020
UR  - http://cherry.chem.bg.ac.rs/handle/123456789/3974
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/3580
AB  - Lignin peroxidase (LiP) is a heme-containing oxidoreductase that oxidizes structurally diverse substrates in an H2O2-dependent manner. Its ability to oxidize many pollutants makes it suitable for bioremediation applications and an ideal candidate for optimization by mutagenesis and selection. In order to increase oxidative stability of LiP we generated a random mutagenesis library comprising 106 mutated LiP genes and screened for expressed enzymes with higher than wild-type activity after incubation in 30 mM H2O2 by flow cytometry with fluorescein-tyramide as a substrate. To preserve the genotype-phenotype connection, the LiP mutants were displayed on the yeast cell surface. Two rounds of sorting were performed, recovered colonies were then screened in microtiter plates, and activity analysis revealed a significant increase in the percentage of cells expressing LiP variants with higher oxidative stability than wtLiP. Two rounds of sorting increased the proportion of more-stable variants from 1.4% in the original library to 52.3%. The most stable variants after two rounds of sorting featured between two and four mutations and retained up to 80% of initial activity after 1 h incubation in 30 mM H2O2. We for the first-time applied flow cytometry for screening of any ligninolytic peroxidase library. Obtained results suggest that developed system may be applied for improvement of industrially important characteristics of lignin peroxidase.
PB  - Elsevier
T2  - Journal of Bioscience and Bioengineering
T1  - Flow cytometry-based system for screening of lignin peroxidase mutants with higher oxidative stability
VL  - 129
IS  - 6
SP  - 664
EP  - 671
DO  - 10.1016/j.jbiosc.2019.12.009
ER  - 

@article{
author = "Ilić Đurđić, Karla and Ece, Selin and Ostafe, Raluca and Vogel, Simon and Balaž, Ana Marija and Schillberg, Stefan and Fischer, Rainer and Prodanović, Radivoje",
year = "2020",
abstract = "Lignin peroxidase (LiP) is a heme-containing oxidoreductase that oxidizes structurally diverse substrates in an H2O2-dependent manner. Its ability to oxidize many pollutants makes it suitable for bioremediation applications and an ideal candidate for optimization by mutagenesis and selection. In order to increase oxidative stability of LiP we generated a random mutagenesis library comprising 106 mutated LiP genes and screened for expressed enzymes with higher than wild-type activity after incubation in 30 mM H2O2 by flow cytometry with fluorescein-tyramide as a substrate. To preserve the genotype-phenotype connection, the LiP mutants were displayed on the yeast cell surface. Two rounds of sorting were performed, recovered colonies were then screened in microtiter plates, and activity analysis revealed a significant increase in the percentage of cells expressing LiP variants with higher oxidative stability than wtLiP. Two rounds of sorting increased the proportion of more-stable variants from 1.4% in the original library to 52.3%. The most stable variants after two rounds of sorting featured between two and four mutations and retained up to 80% of initial activity after 1 h incubation in 30 mM H2O2. We for the first-time applied flow cytometry for screening of any ligninolytic peroxidase library. Obtained results suggest that developed system may be applied for improvement of industrially important characteristics of lignin peroxidase.",
publisher = "Elsevier",
journal = "Journal of Bioscience and Bioengineering",
title = "Flow cytometry-based system for screening of lignin peroxidase mutants with higher oxidative stability",
volume = "129",
number = "6",
pages = "664-671",
doi = "10.1016/j.jbiosc.2019.12.009"
}

Ilić Đurđić, K., Ece, S., Ostafe, R., Vogel, S., Balaž, A. M., Schillberg, S., Fischer, R.,& Prodanović, R.. (2020). Flow cytometry-based system for screening of lignin peroxidase mutants with higher oxidative stability. in Journal of Bioscience and Bioengineering
Elsevier., 129(6), 664-671.
https://doi.org/10.1016/j.jbiosc.2019.12.009

Ilić Đurđić K, Ece S, Ostafe R, Vogel S, Balaž AM, Schillberg S, Fischer R, Prodanović R. Flow cytometry-based system for screening of lignin peroxidase mutants with higher oxidative stability. in Journal of Bioscience and Bioengineering. 2020;129(6):664-671.
doi:10.1016/j.jbiosc.2019.12.009 .

Ilić Đurđić, Karla, Ece, Selin, Ostafe, Raluca, Vogel, Simon, Balaž, Ana Marija, Schillberg, Stefan, Fischer, Rainer, Prodanović, Radivoje, "Flow cytometry-based system for screening of lignin peroxidase mutants with higher oxidative stability" in Journal of Bioscience and Bioengineering, 129, no. 6 (2020):664-671,
https://doi.org/10.1016/j.jbiosc.2019.12.009 . .

TY  - JOUR
AU  - Balaž, Ana Marija
AU  - Blažić, Marija
AU  - Popović, Nikolina
AU  - Prodanović, Olivera
AU  - Ostafe, Raluca
AU  - Fischer, Rainer
AU  - Prodanović, Radivoje
PY  - 2020
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/3880
AB  - Production of soluble cellobiose dehydrogenase (CDH) mutant proteins previously evolved on the surface of S. cerevisiae yeast cells was established for use in biosensors and biofuel cells. For this purpose, mutant cdh genes tm (D20N, A64T, V592M), H5 (D20N, V22A, A64T, V592M) and H9 (D20N, A64T, T84A, A261P, V592M, E674G, N715S) were cloned to pPICZα plasmid and transformed into Pichia pastoris KM71H strain for high expression in a soluble form and kinetic characterization. After 6 days of expression under methanol induction, the CDHs were purified by ultrafiltration, ion- -exchange chromatography and gel filtration. Sodium dodecyl sulfate electrophoresis confirmed the purity and presence of a single protein band at a molecular weight of 100 kDa. Kinetic characterization showed that the H5 mutant had the highest catalytic constant of 43.5 s-1 for lactose, while the mutant H9 showed the highest specificity constant for lactose of 132 mM-1 s-1. All three mutant proteins did not change the pH optimum that was between 4.5 and 5.5. Compared to the previously obtained wild types and mutants of CDH from Phanerochaete chrysosporium, the variants reported in this article had higher activity and specificity that together with high protein expression rate in P. pastoris, makes them good candidates for use in biotechnology for lactobionic acid production and biosensor manufacture.
AB  - У циљу употребе у биосензорима и биогоривним ћелијама, успостављена је производња
растворних облика целобиоза дехидрогеназе (CDH) претходно еволуираних на површини
квашчевих ћелија S. cerevisiae. У ту сврху су мутанти CDH, tm (D20N, A64T, V592M), H5
(D20N, V22A, A64T, V592M) и H9 (D20N, A64T, T84A, A261P, V592M, E674G, N715S)
клонирани у pPICZα плазмид и трансформисани у Pichia pastoris KM71H сој за високу
експресију у растворном облику и кинетичку карактеризацију. После 6 дана експресије под
индукцијом метанолом, мутанти су пречишћени ултрафилтрацијом, јоноизмењивачком
хроматографијом и гел-филтрацијом. SDS електрофореза је потврдила чистоћу уз присуство
једне протеинске траке молекулскe масe од 100 kDa. Кинетичка карактеризација је показала
да H5 мутирани протеин поседује највећу каталитичку константу од 43,5 s-1 за лактозу, док
је H9 имао највећу константу специфичности за лактозу од 132 mM-1 s-1. Сва три мутирана
протеина су имала неизмењен pH оптимум који је био у опсегу од 4,5 до 5,5. У поређењу са
претходно добијеним природним и мутантним облицима CDH протеина из Phanerochaete
chrysosporium, облици приказани у овом раду имају већу активност и специфичност, што их,
повезано са високом експресијом протеина у P. Pastoris, чини добрим кандидатима за упо-
требу у биотехнологији за производњу лактобионске киселине и биосензора.
PB  - Belgrade : Serbian Chemical Society
T2  - Journal of the Serbian Chemical Society
T1  - Expression, purification and characterization of cellobiose dehydrogenase mutants from Phanerochaete chrysosporium in Pichia pastoris KM71H strain
T1  - Ekspresija, prečišćavanje i karakterizacija mutanata celobioza - dehidrogenaze iz Phanerochaete chrysosporium u Pichia pastoris KM71H soju
VL  - 85
IS  - 1
SP  - 25
EP  - 35
DO  - 10.2298/JSC190320058B
ER  - 

@article{
author = "Balaž, Ana Marija and Blažić, Marija and Popović, Nikolina and Prodanović, Olivera and Ostafe, Raluca and Fischer, Rainer and Prodanović, Radivoje",
year = "2020",
abstract = "Production of soluble cellobiose dehydrogenase (CDH) mutant proteins previously evolved on the surface of S. cerevisiae yeast cells was established for use in biosensors and biofuel cells. For this purpose, mutant cdh genes tm (D20N, A64T, V592M), H5 (D20N, V22A, A64T, V592M) and H9 (D20N, A64T, T84A, A261P, V592M, E674G, N715S) were cloned to pPICZα plasmid and transformed into Pichia pastoris KM71H strain for high expression in a soluble form and kinetic characterization. After 6 days of expression under methanol induction, the CDHs were purified by ultrafiltration, ion- -exchange chromatography and gel filtration. Sodium dodecyl sulfate electrophoresis confirmed the purity and presence of a single protein band at a molecular weight of 100 kDa. Kinetic characterization showed that the H5 mutant had the highest catalytic constant of 43.5 s-1 for lactose, while the mutant H9 showed the highest specificity constant for lactose of 132 mM-1 s-1. All three mutant proteins did not change the pH optimum that was between 4.5 and 5.5. Compared to the previously obtained wild types and mutants of CDH from Phanerochaete chrysosporium, the variants reported in this article had higher activity and specificity that together with high protein expression rate in P. pastoris, makes them good candidates for use in biotechnology for lactobionic acid production and biosensor manufacture., У циљу употребе у биосензорима и биогоривним ћелијама, успостављена је производња
растворних облика целобиоза дехидрогеназе (CDH) претходно еволуираних на површини
квашчевих ћелија S. cerevisiae. У ту сврху су мутанти CDH, tm (D20N, A64T, V592M), H5
(D20N, V22A, A64T, V592M) и H9 (D20N, A64T, T84A, A261P, V592M, E674G, N715S)
клонирани у pPICZα плазмид и трансформисани у Pichia pastoris KM71H сој за високу
експресију у растворном облику и кинетичку карактеризацију. После 6 дана експресије под
индукцијом метанолом, мутанти су пречишћени ултрафилтрацијом, јоноизмењивачком
хроматографијом и гел-филтрацијом. SDS електрофореза је потврдила чистоћу уз присуство
једне протеинске траке молекулскe масe од 100 kDa. Кинетичка карактеризација је показала
да H5 мутирани протеин поседује највећу каталитичку константу од 43,5 s-1 за лактозу, док
је H9 имао највећу константу специфичности за лактозу од 132 mM-1 s-1. Сва три мутирана
протеина су имала неизмењен pH оптимум који је био у опсегу од 4,5 до 5,5. У поређењу са
претходно добијеним природним и мутантним облицима CDH протеина из Phanerochaete
chrysosporium, облици приказани у овом раду имају већу активност и специфичност, што их,
повезано са високом експресијом протеина у P. Pastoris, чини добрим кандидатима за упо-
требу у биотехнологији за производњу лактобионске киселине и биосензора.",
publisher = "Belgrade : Serbian Chemical Society",
journal = "Journal of the Serbian Chemical Society",
title = "Expression, purification and characterization of cellobiose dehydrogenase mutants from Phanerochaete chrysosporium in Pichia pastoris KM71H strain, Ekspresija, prečišćavanje i karakterizacija mutanata celobioza - dehidrogenaze iz Phanerochaete chrysosporium u Pichia pastoris KM71H soju",
volume = "85",
number = "1",
pages = "25-35",
doi = "10.2298/JSC190320058B"
}

Balaž, A. M., Blažić, M., Popović, N., Prodanović, O., Ostafe, R., Fischer, R.,& Prodanović, R.. (2020). Expression, purification and characterization of cellobiose dehydrogenase mutants from Phanerochaete chrysosporium in Pichia pastoris KM71H strain. in Journal of the Serbian Chemical Society
Belgrade : Serbian Chemical Society., 85(1), 25-35.
https://doi.org/10.2298/JSC190320058B

Balaž AM, Blažić M, Popović N, Prodanović O, Ostafe R, Fischer R, Prodanović R. Expression, purification and characterization of cellobiose dehydrogenase mutants from Phanerochaete chrysosporium in Pichia pastoris KM71H strain. in Journal of the Serbian Chemical Society. 2020;85(1):25-35.
doi:10.2298/JSC190320058B .

Balaž, Ana Marija, Blažić, Marija, Popović, Nikolina, Prodanović, Olivera, Ostafe, Raluca, Fischer, Rainer, Prodanović, Radivoje, "Expression, purification and characterization of cellobiose dehydrogenase mutants from Phanerochaete chrysosporium in Pichia pastoris KM71H strain" in Journal of the Serbian Chemical Society, 85, no. 1 (2020):25-35,
https://doi.org/10.2298/JSC190320058B . .

TY  - CONF
AU  - Milošević, Jelica
AU  - Mijin, Nemanja
AU  - Maleš, Luka
AU  - Margetić, Aleksandra
AU  - Jovčić, Branko
AU  - Polović, Natalija
PY  - 2019
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/5817
AB  - Proteins are prone to structural changes due to their marginal stability. There are multiple
pathways of structural rearrangements leading to misfolding and aggregation among which
amyloids stand out as highly ordered and remarkably stable forms which appear to be a
global minimum of protein free energy landscape of all proteins. In vitro studies on
different proteins show that destabilizing conditions that favor the state of molten globule
are likely to lead to ordered fibril formation. The presence of various organic and inorganic
molecules was reported to affect amyloid fibril formation, eighter as stimulators or
inhibitors. We investigated the formation of amyloid fibrils of human serum albumin,
ovalbumin and papain in the presence of metal ions, as well as low-molecular-weight
compounds. Proteins were incubated in destabilizing conditions optimized to prolong the
solubility of molten globule state and induce amyloid-like structural changes. The effects
of inorganic and organic additives on fibrillation process were monitored using Thioflavin
T fluorescence, 8-Anilinonaphthalene-1-sulfonic acid fluorescence, Attenuated total
reflection Fourier-transform infrared spectroscopy, electrophoretic and microscopy
techniques. Our results show that the kinetics of amyloid formation is dependent on the
presence of iron, copper, zinc and aluminum salts, as well as different lipophilic lowmolecular-
weight compounds. While some compounds act as complete inhibitors of
fibrillation, others increase the rate of fibrillation process and promote the formation of
mature fibrils.
PB  - University of Belgrade - Falulty of chemistry
PB  - Serbian Biochemical Society
C3  - Proceedings - IX Conference of Serbian Biochemical Society “Diversity in Biochemistry”, 14-16.11.2019. Belgrade, Serbia
T1  - Kinetics of amyloid fibril formation in the presence of metal ions and low-molecular-weight compounds
SP  - 133
UR  - https://hdl.handle.net/21.15107/rcub_cer_5817
ER  - 

@conference{
author = "Milošević, Jelica and Mijin, Nemanja and Maleš, Luka and Margetić, Aleksandra and Jovčić, Branko and Polović, Natalija",
year = "2019",
abstract = "Proteins are prone to structural changes due to their marginal stability. There are multiple
pathways of structural rearrangements leading to misfolding and aggregation among which
amyloids stand out as highly ordered and remarkably stable forms which appear to be a
global minimum of protein free energy landscape of all proteins. In vitro studies on
different proteins show that destabilizing conditions that favor the state of molten globule
are likely to lead to ordered fibril formation. The presence of various organic and inorganic
molecules was reported to affect amyloid fibril formation, eighter as stimulators or
inhibitors. We investigated the formation of amyloid fibrils of human serum albumin,
ovalbumin and papain in the presence of metal ions, as well as low-molecular-weight
compounds. Proteins were incubated in destabilizing conditions optimized to prolong the
solubility of molten globule state and induce amyloid-like structural changes. The effects
of inorganic and organic additives on fibrillation process were monitored using Thioflavin
T fluorescence, 8-Anilinonaphthalene-1-sulfonic acid fluorescence, Attenuated total
reflection Fourier-transform infrared spectroscopy, electrophoretic and microscopy
techniques. Our results show that the kinetics of amyloid formation is dependent on the
presence of iron, copper, zinc and aluminum salts, as well as different lipophilic lowmolecular-
weight compounds. While some compounds act as complete inhibitors of
fibrillation, others increase the rate of fibrillation process and promote the formation of
mature fibrils.",
publisher = "University of Belgrade - Falulty of chemistry, Serbian Biochemical Society",
journal = "Proceedings - IX Conference of Serbian Biochemical Society “Diversity in Biochemistry”, 14-16.11.2019. Belgrade, Serbia",
title = "Kinetics of amyloid fibril formation in the presence of metal ions and low-molecular-weight compounds",
pages = "133",
url = "https://hdl.handle.net/21.15107/rcub_cer_5817"
}

Milošević, J., Mijin, N., Maleš, L., Margetić, A., Jovčić, B.,& Polović, N.. (2019). Kinetics of amyloid fibril formation in the presence of metal ions and low-molecular-weight compounds. in Proceedings - IX Conference of Serbian Biochemical Society “Diversity in Biochemistry”, 14-16.11.2019. Belgrade, Serbia
University of Belgrade - Falulty of chemistry., 133.
https://hdl.handle.net/21.15107/rcub_cer_5817

Milošević J, Mijin N, Maleš L, Margetić A, Jovčić B, Polović N. Kinetics of amyloid fibril formation in the presence of metal ions and low-molecular-weight compounds. in Proceedings - IX Conference of Serbian Biochemical Society “Diversity in Biochemistry”, 14-16.11.2019. Belgrade, Serbia. 2019;:133.
https://hdl.handle.net/21.15107/rcub_cer_5817 .

Milošević, Jelica, Mijin, Nemanja, Maleš, Luka, Margetić, Aleksandra, Jovčić, Branko, Polović, Natalija, "Kinetics of amyloid fibril formation in the presence of metal ions and low-molecular-weight compounds" in Proceedings - IX Conference of Serbian Biochemical Society “Diversity in Biochemistry”, 14-16.11.2019. Belgrade, Serbia (2019):133,
https://hdl.handle.net/21.15107/rcub_cer_5817 .

TY  - JOUR
AU  - Spasojević, Dragica
AU  - Prokopijević, Miloš
AU  - Prodanović, Olivera
AU  - Zelenović, Nevena
AU  - Polović, Natalija
AU  - Radotić, Ksenija
AU  - Prodanović, Radivoje
PY  - 2019
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/3028
AB  - Derivatives of xylans were synthesized from corncob xylan by carboxymethylation, oxidization with different molar ratios of periodate (5, 10 15 and 20 mol%) and by reductive amination with tyramine. Modifications of tyramine carboxymethyl xylans (Tyr-CMX) were confirmed by FTIR, UV and NMR spectra. Concentration of ionizable groups increased from 1.5 mmol/g for carboxymethyl xylan (CMX) to 5.4 mmol/g for Tyr-CMX oxidized with 20 mol% of periodate. All Tyr-CMXs were able to form hydrogels the cross-linking reaction with horseradish peroxidase and peroxide. Tyr-CMXs were tested for amyloglucosidase (AG) encapsulation within hydrogel microbeads obtained in a reaction of emulsion polymerization with peroxidase. Average diameter of Tyr-CMX hydrogel microbeads was 52±25 µm and after encapsulation optimization with respect to the extent of CMX modification with tyramine, the concentration of Tyr-CMX, and the amount of added AG, microbeads with AG specific activity of 2 U/mL and 20% yield of immobilization were obtained. The optimum pH of the immobilized AG was not changed compared to the soluble one, while half-life at 60 °C was increased around 10 times. The Michaelis-Menten constant for the immobilized enzyme, 1.03 mM, was significantly lower than that for the soluble one, 1.54 mM. After 5 cycles of repetitive use in batch reactor, the immobilized AG retained 68% of initial activity.
PB  - Springer
T2  - Macromolecular Research
T1  - Peroxidase-Sensitive Tyramine Carboxymethyl Xylan Hydrogels for Enzyme Encapsulation
VL  - 27
IS  - 8
SP  - 764
EP  - 771
DO  - 10.1007/s13233-019-7111-7
ER  - 

@article{
author = "Spasojević, Dragica and Prokopijević, Miloš and Prodanović, Olivera and Zelenović, Nevena and Polović, Natalija and Radotić, Ksenija and Prodanović, Radivoje",
year = "2019",
abstract = "Derivatives of xylans were synthesized from corncob xylan by carboxymethylation, oxidization with different molar ratios of periodate (5, 10 15 and 20 mol%) and by reductive amination with tyramine. Modifications of tyramine carboxymethyl xylans (Tyr-CMX) were confirmed by FTIR, UV and NMR spectra. Concentration of ionizable groups increased from 1.5 mmol/g for carboxymethyl xylan (CMX) to 5.4 mmol/g for Tyr-CMX oxidized with 20 mol% of periodate. All Tyr-CMXs were able to form hydrogels the cross-linking reaction with horseradish peroxidase and peroxide. Tyr-CMXs were tested for amyloglucosidase (AG) encapsulation within hydrogel microbeads obtained in a reaction of emulsion polymerization with peroxidase. Average diameter of Tyr-CMX hydrogel microbeads was 52±25 µm and after encapsulation optimization with respect to the extent of CMX modification with tyramine, the concentration of Tyr-CMX, and the amount of added AG, microbeads with AG specific activity of 2 U/mL and 20% yield of immobilization were obtained. The optimum pH of the immobilized AG was not changed compared to the soluble one, while half-life at 60 °C was increased around 10 times. The Michaelis-Menten constant for the immobilized enzyme, 1.03 mM, was significantly lower than that for the soluble one, 1.54 mM. After 5 cycles of repetitive use in batch reactor, the immobilized AG retained 68% of initial activity.",
publisher = "Springer",
journal = "Macromolecular Research",
title = "Peroxidase-Sensitive Tyramine Carboxymethyl Xylan Hydrogels for Enzyme Encapsulation",
volume = "27",
number = "8",
pages = "764-771",
doi = "10.1007/s13233-019-7111-7"
}

Spasojević, D., Prokopijević, M., Prodanović, O., Zelenović, N., Polović, N., Radotić, K.,& Prodanović, R.. (2019). Peroxidase-Sensitive Tyramine Carboxymethyl Xylan Hydrogels for Enzyme Encapsulation. in Macromolecular Research
Springer., 27(8), 764-771.
https://doi.org/10.1007/s13233-019-7111-7

Spasojević D, Prokopijević M, Prodanović O, Zelenović N, Polović N, Radotić K, Prodanović R. Peroxidase-Sensitive Tyramine Carboxymethyl Xylan Hydrogels for Enzyme Encapsulation. in Macromolecular Research. 2019;27(8):764-771.
doi:10.1007/s13233-019-7111-7 .

Spasojević, Dragica, Prokopijević, Miloš, Prodanović, Olivera, Zelenović, Nevena, Polović, Natalija, Radotić, Ksenija, Prodanović, Radivoje, "Peroxidase-Sensitive Tyramine Carboxymethyl Xylan Hydrogels for Enzyme Encapsulation" in Macromolecular Research, 27, no. 8 (2019):764-771,
https://doi.org/10.1007/s13233-019-7111-7 . .

TY  - JOUR
AU  - Uzelac, Tamara N.
AU  - Nikolić-Kokić, Aleksandra
AU  - Spasić, Snežana
AU  - Mačvanin, Mirjana T.
AU  - Nikolić, Milan
AU  - Mandić, Ljuba M.
AU  - Jovanović, Vesna B.
PY  - 2019
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/3432
AB  - Antipsychotic drugs interfere with the antioxidant defense system provoking complex and often toxicological effects. Here we examined differences in plasma albumin reduced free thiol (SH) group content and its reactivity as a consequence of clozapine (CLZ) and ziprasidone (ZIP) binding. Chronic administration of CLZ reduced, whereas treatment with ZIP increased albumin-SH content in rats. Regardless of the ratio of stearic acid (SA) bound to protein, in vitro binding of ZIP to human serum albumin (HSA) increased both the SH group level and reactivity. In contrast, the effect of CLZ on HSA-SH reactivity was dependent on HSA to SA molar ratio. CLZ binding was accompanied by an increase in HSA-SH reactivity in samples with normal, but a reduction of itsreactivity level with higher SA/HSA ratio, compared to drug-free samples. We demonstrate by steady-state fluorescence quenching studies that an increase in SA binding to HSA is associated with a significant reduction of binding constant for both antipsychotics. In addition, this is the first report of quantitative characterization of ZIP binding to HSA. Our findings suggest that albumin-SH content and reactivity is modulated by ZIP towards an increased antioxidant defense capacity in circulation, as opposed to CLZ, which can contribute to the safer, more effective treatment of schizophrenia.
PB  - Elsevier
T2  - Chemico-Biological Interactions
T1  - Opposite clozapine and ziprasidone effects on the reactivity of plasma albumin SH-group are the consequence of their different binding properties dependent on protein fatty acids content
VL  - 311
SP  - 108787
DO  - 10.1016/j.cbi.2019.108787
ER  - 

@article{
author = "Uzelac, Tamara N. and Nikolić-Kokić, Aleksandra and Spasić, Snežana and Mačvanin, Mirjana T. and Nikolić, Milan and Mandić, Ljuba M. and Jovanović, Vesna B.",
year = "2019",
abstract = "Antipsychotic drugs interfere with the antioxidant defense system provoking complex and often toxicological effects. Here we examined differences in plasma albumin reduced free thiol (SH) group content and its reactivity as a consequence of clozapine (CLZ) and ziprasidone (ZIP) binding. Chronic administration of CLZ reduced, whereas treatment with ZIP increased albumin-SH content in rats. Regardless of the ratio of stearic acid (SA) bound to protein, in vitro binding of ZIP to human serum albumin (HSA) increased both the SH group level and reactivity. In contrast, the effect of CLZ on HSA-SH reactivity was dependent on HSA to SA molar ratio. CLZ binding was accompanied by an increase in HSA-SH reactivity in samples with normal, but a reduction of itsreactivity level with higher SA/HSA ratio, compared to drug-free samples. We demonstrate by steady-state fluorescence quenching studies that an increase in SA binding to HSA is associated with a significant reduction of binding constant for both antipsychotics. In addition, this is the first report of quantitative characterization of ZIP binding to HSA. Our findings suggest that albumin-SH content and reactivity is modulated by ZIP towards an increased antioxidant defense capacity in circulation, as opposed to CLZ, which can contribute to the safer, more effective treatment of schizophrenia.",
publisher = "Elsevier",
journal = "Chemico-Biological Interactions",
title = "Opposite clozapine and ziprasidone effects on the reactivity of plasma albumin SH-group are the consequence of their different binding properties dependent on protein fatty acids content",
volume = "311",
pages = "108787",
doi = "10.1016/j.cbi.2019.108787"
}

Uzelac, T. N., Nikolić-Kokić, A., Spasić, S., Mačvanin, M. T., Nikolić, M., Mandić, L. M.,& Jovanović, V. B.. (2019). Opposite clozapine and ziprasidone effects on the reactivity of plasma albumin SH-group are the consequence of their different binding properties dependent on protein fatty acids content. in Chemico-Biological Interactions
Elsevier., 311, 108787.
https://doi.org/10.1016/j.cbi.2019.108787

Uzelac TN, Nikolić-Kokić A, Spasić S, Mačvanin MT, Nikolić M, Mandić LM, Jovanović VB. Opposite clozapine and ziprasidone effects on the reactivity of plasma albumin SH-group are the consequence of their different binding properties dependent on protein fatty acids content. in Chemico-Biological Interactions. 2019;311:108787.
doi:10.1016/j.cbi.2019.108787 .

Uzelac, Tamara N., Nikolić-Kokić, Aleksandra, Spasić, Snežana, Mačvanin, Mirjana T., Nikolić, Milan, Mandić, Ljuba M., Jovanović, Vesna B., "Opposite clozapine and ziprasidone effects on the reactivity of plasma albumin SH-group are the consequence of their different binding properties dependent on protein fatty acids content" in Chemico-Biological Interactions, 311 (2019):108787,
https://doi.org/10.1016/j.cbi.2019.108787 . .

TY  - JOUR
AU  - Uzelac, Tamara N.
AU  - Nikolić-Kokić, Aleksandra
AU  - Spasić, Snežana
AU  - Mačvanin, Mirjana T.
AU  - Nikolić, Milan
AU  - Mandić, Ljuba M.
AU  - Jovanović, Vesna B.
PY  - 2019
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/3053
AB  - Antipsychotic drugs interfere with the antioxidant defense system provoking complex and often toxicological effects. Here we examined differences in plasma albumin reduced free thiol (SH) group content and its reactivity as a consequence of clozapine (CLZ) and ziprasidone (ZIP) binding. Chronic administration of CLZ reduced, whereas treatment with ZIP increased albumin-SH content in rats. Regardless of the ratio of stearic acid (SA) bound to protein, in vitro binding of ZIP to human serum albumin (HSA) increased both the SH group level and reactivity. In contrast, the effect of CLZ on HSA-SH reactivity was dependent on HSA to SA molar ratio. CLZ binding was accompanied by an increase in HSA-SH reactivity in samples with normal, but a reduction of its
reactivity level with higher SA/HSA ratio, compared to drug-free samples. We demonstrate by steady-state fluorescence quenching studies that an increase in SA binding to HSA is associated with a significant reduction of binding constant for both antipsychotics. In addition, this is the first report of quantitative characterization of ZIP binding to HSA. Our findings suggest that albumin-SH content and reactivity is modulated by ZIP towards an increased antioxidant defense capacity in circulation, as opposed to CLZ, which can contribute to the safer, more effective treatment of schizophrenia.
PB  - Elsevier
T2  - Chemico-Biological Interactions
T1  - Opposite clozapine and ziprasidone effects on the reactivity of plasma albumin SH-group are the consequence of their different binding properties dependent on protein fatty acids content
VL  - 311
SP  - 108787
DO  - 10.1016/j.cbi.2019.108787
ER  - 

@article{
author = "Uzelac, Tamara N. and Nikolić-Kokić, Aleksandra and Spasić, Snežana and Mačvanin, Mirjana T. and Nikolić, Milan and Mandić, Ljuba M. and Jovanović, Vesna B.",
year = "2019",
abstract = "Antipsychotic drugs interfere with the antioxidant defense system provoking complex and often toxicological effects. Here we examined differences in plasma albumin reduced free thiol (SH) group content and its reactivity as a consequence of clozapine (CLZ) and ziprasidone (ZIP) binding. Chronic administration of CLZ reduced, whereas treatment with ZIP increased albumin-SH content in rats. Regardless of the ratio of stearic acid (SA) bound to protein, in vitro binding of ZIP to human serum albumin (HSA) increased both the SH group level and reactivity. In contrast, the effect of CLZ on HSA-SH reactivity was dependent on HSA to SA molar ratio. CLZ binding was accompanied by an increase in HSA-SH reactivity in samples with normal, but a reduction of its
reactivity level with higher SA/HSA ratio, compared to drug-free samples. We demonstrate by steady-state fluorescence quenching studies that an increase in SA binding to HSA is associated with a significant reduction of binding constant for both antipsychotics. In addition, this is the first report of quantitative characterization of ZIP binding to HSA. Our findings suggest that albumin-SH content and reactivity is modulated by ZIP towards an increased antioxidant defense capacity in circulation, as opposed to CLZ, which can contribute to the safer, more effective treatment of schizophrenia.",
publisher = "Elsevier",
journal = "Chemico-Biological Interactions",
title = "Opposite clozapine and ziprasidone effects on the reactivity of plasma albumin SH-group are the consequence of their different binding properties dependent on protein fatty acids content",
volume = "311",
pages = "108787",
doi = "10.1016/j.cbi.2019.108787"
}

Uzelac, T. N., Nikolić-Kokić, A., Spasić, S., Mačvanin, M. T., Nikolić, M., Mandić, L. M.,& Jovanović, V. B.. (2019). Opposite clozapine and ziprasidone effects on the reactivity of plasma albumin SH-group are the consequence of their different binding properties dependent on protein fatty acids content. in Chemico-Biological Interactions
Elsevier., 311, 108787.
https://doi.org/10.1016/j.cbi.2019.108787

Uzelac TN, Nikolić-Kokić A, Spasić S, Mačvanin MT, Nikolić M, Mandić LM, Jovanović VB. Opposite clozapine and ziprasidone effects on the reactivity of plasma albumin SH-group are the consequence of their different binding properties dependent on protein fatty acids content. in Chemico-Biological Interactions. 2019;311:108787.
doi:10.1016/j.cbi.2019.108787 .

Uzelac, Tamara N., Nikolić-Kokić, Aleksandra, Spasić, Snežana, Mačvanin, Mirjana T., Nikolić, Milan, Mandić, Ljuba M., Jovanović, Vesna B., "Opposite clozapine and ziprasidone effects on the reactivity of plasma albumin SH-group are the consequence of their different binding properties dependent on protein fatty acids content" in Chemico-Biological Interactions, 311 (2019):108787,
https://doi.org/10.1016/j.cbi.2019.108787 . .

TY  - JOUR
AU  - Kovačević, Gordana
AU  - Ostafe, Raluca
AU  - Balaž, Ana Marija
AU  - Fischer, Rainer
AU  - Prodanović, Radivoje
PY  - 2019
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/2497
AB  - Glucose oxidase (GOx) mutants with higher activity or stability have important role in industry and in the development of biosensors and biofuel cells. Discovering these mutants can be time-consuming if appropriate high-throughput screening (HTS) systems are not available. GOx gene libraries were successfully screened and sorted using a HTS system based on GOx activity dependent fluorescent labeling of yeast cells with tyramids and quantification of the amount of expressed enzyme by yeast enhanced green fluorescent protein (yGFP) tagging and flow cytometry. For this purpose, we expressed wild type and a mutant GOx as a chimera with the yGFP to confirm differences in catalytic activity between wild-type and mutant GOx. Fluorescence of yGFP is preserved during expression of chimera, and also after the oxidative enzymatic reaction. We have obtained a 2.5-fold enrichment in population of cells expressing active enzyme, and percentage of enzyme variants with enzymatic mean activity higher than wild type activity was increased to 44% after a single round of GOx gene library sorting. We have found two mutants with 1.3 and 2.3-fold increase in Vmax values compared to the wtGOx. By simultaneous detection of protein expression level and enzyme activity we have increased the likelihood of finding GOx variants with increased activity in a single round of flow cytometry sorting.
PB  - Elsevier
T2  - Journal of Bioscience and Bioengineering
T1  - Development of GFP-based high-throughput screening system for directed evolution of glucose oxidase
VL  - 127
IS  - 1
SP  - 30
EP  - 37
DO  - 10.1016/j.jbiosc.2018.07.002
ER  - 

@article{
author = "Kovačević, Gordana and Ostafe, Raluca and Balaž, Ana Marija and Fischer, Rainer and Prodanović, Radivoje",
year = "2019",
abstract = "Glucose oxidase (GOx) mutants with higher activity or stability have important role in industry and in the development of biosensors and biofuel cells. Discovering these mutants can be time-consuming if appropriate high-throughput screening (HTS) systems are not available. GOx gene libraries were successfully screened and sorted using a HTS system based on GOx activity dependent fluorescent labeling of yeast cells with tyramids and quantification of the amount of expressed enzyme by yeast enhanced green fluorescent protein (yGFP) tagging and flow cytometry. For this purpose, we expressed wild type and a mutant GOx as a chimera with the yGFP to confirm differences in catalytic activity between wild-type and mutant GOx. Fluorescence of yGFP is preserved during expression of chimera, and also after the oxidative enzymatic reaction. We have obtained a 2.5-fold enrichment in population of cells expressing active enzyme, and percentage of enzyme variants with enzymatic mean activity higher than wild type activity was increased to 44% after a single round of GOx gene library sorting. We have found two mutants with 1.3 and 2.3-fold increase in Vmax values compared to the wtGOx. By simultaneous detection of protein expression level and enzyme activity we have increased the likelihood of finding GOx variants with increased activity in a single round of flow cytometry sorting.",
publisher = "Elsevier",
journal = "Journal of Bioscience and Bioengineering",
title = "Development of GFP-based high-throughput screening system for directed evolution of glucose oxidase",
volume = "127",
number = "1",
pages = "30-37",
doi = "10.1016/j.jbiosc.2018.07.002"
}

Kovačević, G., Ostafe, R., Balaž, A. M., Fischer, R.,& Prodanović, R.. (2019). Development of GFP-based high-throughput screening system for directed evolution of glucose oxidase. in Journal of Bioscience and Bioengineering
Elsevier., 127(1), 30-37.
https://doi.org/10.1016/j.jbiosc.2018.07.002

Kovačević G, Ostafe R, Balaž AM, Fischer R, Prodanović R. Development of GFP-based high-throughput screening system for directed evolution of glucose oxidase. in Journal of Bioscience and Bioengineering. 2019;127(1):30-37.
doi:10.1016/j.jbiosc.2018.07.002 .

Kovačević, Gordana, Ostafe, Raluca, Balaž, Ana Marija, Fischer, Rainer, Prodanović, Radivoje, "Development of GFP-based high-throughput screening system for directed evolution of glucose oxidase" in Journal of Bioscience and Bioengineering, 127, no. 1 (2019):30-37,
https://doi.org/10.1016/j.jbiosc.2018.07.002 . .

TY  - JOUR
AU  - Blažić, Marija
AU  - Balaž, Ana Marija
AU  - Tadić, Vojin
AU  - Draganić, Bojana
AU  - Ostafe, Raluca
AU  - Fischer, Rainer
AU  - Prodanović, Radivoje
PY  - 2019
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/2655
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/2656
AB  - Cellobiose dehydrogenase (CDH) can be used in industry for lactobionic acid production, as a part of biosensors for disaccharides and in wound healing. In fungi it is involved in lignocellulose degradation. CDH gene from Phanerochaete chrysosporium has been cloned in pYES2 plasmid for extracellular expression and protein engineering in yeast Saccharomyces cerevisiae InvSC1 for the first time. A CDH gene library was generated using error-prone PCR and screened by spectrophotometric enzymatic assaybased on 2,6-dichloroindophenol reduction detection in microtiter plates. Several mutants with increased activity and specificity towards lactose and cellobiose were found, purified and characterized in detail. Recombinant CDH enzymes showed a broad molecular weight between 120 and 150 KDa due to hyperglycosylation and the best S137N mutant showed 2.2 times increased kcat and 1.5 and 2 times increased specificity constant for lactose and cellobiose compared to the wild type enzyme. pH optimum of mutants was not changed while thermostability of selected mutants improved and S137N mutant retained 30% of it’s original activity after 15 minutes at 70oC compared to 10% of activity that the wild type enzyme retained. Mutants M65S and S137N showed also 1.6 and 1.5 times increased productivity of hydrogen peroxide in the presence of 30mM lactose compared to the wild type.
PB  - Elsevier
T2  - Biochemical Engineering Journal
T1  - Protein engineering of cellobiose dehydrogenase from Phanerochaete chrysosporium in yeast Saccharomyces cerevisiae InvSc1 for increased activity and stability
VL  - 146
SP  - 179
EP  - 185
DO  - 10.1016/j.bej.2019.03.025
ER  - 

@article{
author = "Blažić, Marija and Balaž, Ana Marija and Tadić, Vojin and Draganić, Bojana and Ostafe, Raluca and Fischer, Rainer and Prodanović, Radivoje",
year = "2019",
abstract = "Cellobiose dehydrogenase (CDH) can be used in industry for lactobionic acid production, as a part of biosensors for disaccharides and in wound healing. In fungi it is involved in lignocellulose degradation. CDH gene from Phanerochaete chrysosporium has been cloned in pYES2 plasmid for extracellular expression and protein engineering in yeast Saccharomyces cerevisiae InvSC1 for the first time. A CDH gene library was generated using error-prone PCR and screened by spectrophotometric enzymatic assaybased on 2,6-dichloroindophenol reduction detection in microtiter plates. Several mutants with increased activity and specificity towards lactose and cellobiose were found, purified and characterized in detail. Recombinant CDH enzymes showed a broad molecular weight between 120 and 150 KDa due to hyperglycosylation and the best S137N mutant showed 2.2 times increased kcat and 1.5 and 2 times increased specificity constant for lactose and cellobiose compared to the wild type enzyme. pH optimum of mutants was not changed while thermostability of selected mutants improved and S137N mutant retained 30% of it’s original activity after 15 minutes at 70oC compared to 10% of activity that the wild type enzyme retained. Mutants M65S and S137N showed also 1.6 and 1.5 times increased productivity of hydrogen peroxide in the presence of 30mM lactose compared to the wild type.",
publisher = "Elsevier",
journal = "Biochemical Engineering Journal",
title = "Protein engineering of cellobiose dehydrogenase from Phanerochaete chrysosporium in yeast Saccharomyces cerevisiae InvSc1 for increased activity and stability",
volume = "146",
pages = "179-185",
doi = "10.1016/j.bej.2019.03.025"
}

Blažić, M., Balaž, A. M., Tadić, V., Draganić, B., Ostafe, R., Fischer, R.,& Prodanović, R.. (2019). Protein engineering of cellobiose dehydrogenase from Phanerochaete chrysosporium in yeast Saccharomyces cerevisiae InvSc1 for increased activity and stability. in Biochemical Engineering Journal
Elsevier., 146, 179-185.
https://doi.org/10.1016/j.bej.2019.03.025

Blažić M, Balaž AM, Tadić V, Draganić B, Ostafe R, Fischer R, Prodanović R. Protein engineering of cellobiose dehydrogenase from Phanerochaete chrysosporium in yeast Saccharomyces cerevisiae InvSc1 for increased activity and stability. in Biochemical Engineering Journal. 2019;146:179-185.
doi:10.1016/j.bej.2019.03.025 .

Blažić, Marija, Balaž, Ana Marija, Tadić, Vojin, Draganić, Bojana, Ostafe, Raluca, Fischer, Rainer, Prodanović, Radivoje, "Protein engineering of cellobiose dehydrogenase from Phanerochaete chrysosporium in yeast Saccharomyces cerevisiae InvSc1 for increased activity and stability" in Biochemical Engineering Journal, 146 (2019):179-185,
https://doi.org/10.1016/j.bej.2019.03.025 . .

TY  - JOUR
AU  - Blažić, Marija
AU  - Balaž, Ana Marija
AU  - Tadić, Vojin
AU  - Draganić, Bojana
AU  - Ostafe, Raluca
AU  - Fischer, Rainer
AU  - Prodanović, Radivoje
PY  - 2019
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/2655
AB  - Cellobiose dehydrogenase (CDH) can be used in industry for lactobionic acid production, as a part of biosensors for disaccharides and in wound healing. In fungi it is involved in lignocellulose degradation. CDH gene from Phanerochaete chrysosporium has been cloned in pYES2 plasmid for extracellular expression and protein engineering in yeast Saccharomyces cerevisiae InvSC1 for the first time. A CDH gene library was generated using error-prone PCR and screened by spectrophotometric enzymatic assay
based on 2,6-dichloroindophenol reduction detection in microtiter plates. Several mutants with increased activity and specificity towards lactose and cellobiose were found, purified and characterized in detail. Recombinant CDH enzymes showed a broad molecular weight between 120 and 150 KDa due to hyperglycosylation and the best S137N mutant showed 2.2 times increased kcat and 1.5 and 2 times increased specificity constant for lactose and cellobiose compared to the wild type enzyme. pH optimum of mutants was not changed while thermostability of selected mutants improved and S137N mutant retained 30% of it’s original activity after 15 minutes at 70oC compared to 10% of activity that the wild type enzyme retained. Mutants M65S and S137N showed also 1.6 and 1.5 times increased productivity of hydrogen peroxide in the presence of 30mM lactose compared to the wild type.
PB  - Elsevier
T2  - Biochemical Engineering Journal
T1  - Protein engineering of cellobiose dehydrogenase from Phanerochaete chrysosporium in yeast Saccharomyces cerevisiae InvSc1 for increased activity and stability
VL  - 146
SP  - 179
EP  - 185
DO  - 10.1016/j.bej.2019.03.025
ER  - 

@article{
author = "Blažić, Marija and Balaž, Ana Marija and Tadić, Vojin and Draganić, Bojana and Ostafe, Raluca and Fischer, Rainer and Prodanović, Radivoje",
year = "2019",
abstract = "Cellobiose dehydrogenase (CDH) can be used in industry for lactobionic acid production, as a part of biosensors for disaccharides and in wound healing. In fungi it is involved in lignocellulose degradation. CDH gene from Phanerochaete chrysosporium has been cloned in pYES2 plasmid for extracellular expression and protein engineering in yeast Saccharomyces cerevisiae InvSC1 for the first time. A CDH gene library was generated using error-prone PCR and screened by spectrophotometric enzymatic assay
based on 2,6-dichloroindophenol reduction detection in microtiter plates. Several mutants with increased activity and specificity towards lactose and cellobiose were found, purified and characterized in detail. Recombinant CDH enzymes showed a broad molecular weight between 120 and 150 KDa due to hyperglycosylation and the best S137N mutant showed 2.2 times increased kcat and 1.5 and 2 times increased specificity constant for lactose and cellobiose compared to the wild type enzyme. pH optimum of mutants was not changed while thermostability of selected mutants improved and S137N mutant retained 30% of it’s original activity after 15 minutes at 70oC compared to 10% of activity that the wild type enzyme retained. Mutants M65S and S137N showed also 1.6 and 1.5 times increased productivity of hydrogen peroxide in the presence of 30mM lactose compared to the wild type.",
publisher = "Elsevier",
journal = "Biochemical Engineering Journal",
title = "Protein engineering of cellobiose dehydrogenase from Phanerochaete chrysosporium in yeast Saccharomyces cerevisiae InvSc1 for increased activity and stability",
volume = "146",
pages = "179-185",
doi = "10.1016/j.bej.2019.03.025"
}

Blažić, M., Balaž, A. M., Tadić, V., Draganić, B., Ostafe, R., Fischer, R.,& Prodanović, R.. (2019). Protein engineering of cellobiose dehydrogenase from Phanerochaete chrysosporium in yeast Saccharomyces cerevisiae InvSc1 for increased activity and stability. in Biochemical Engineering Journal
Elsevier., 146, 179-185.
https://doi.org/10.1016/j.bej.2019.03.025

Blažić M, Balaž AM, Tadić V, Draganić B, Ostafe R, Fischer R, Prodanović R. Protein engineering of cellobiose dehydrogenase from Phanerochaete chrysosporium in yeast Saccharomyces cerevisiae InvSc1 for increased activity and stability. in Biochemical Engineering Journal. 2019;146:179-185.
doi:10.1016/j.bej.2019.03.025 .

Blažić, Marija, Balaž, Ana Marija, Tadić, Vojin, Draganić, Bojana, Ostafe, Raluca, Fischer, Rainer, Prodanović, Radivoje, "Protein engineering of cellobiose dehydrogenase from Phanerochaete chrysosporium in yeast Saccharomyces cerevisiae InvSc1 for increased activity and stability" in Biochemical Engineering Journal, 146 (2019):179-185,
https://doi.org/10.1016/j.bej.2019.03.025 . .

TY  - JOUR
AU  - Mrkić, Ivan
AU  - Minić, Rajna
AU  - Popović, Dragan
AU  - Zivkovic, Irena
AU  - Gavrović-Jankulović, Marija
PY  - 2018
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/2380
AB  - Aim To investigate the immunomodulatory potential of a chimera composed of the receptor-binding domain of hemagglutinin 1 (H1s) from Influenza virus and Der p 2 (D2) allergen for allergen-specific immunotherapy of house-dust mite allergy (HDM). Main methods: H1sD2 chimera and D2 allergen were produced by genetic engineering in E. coli. Recombinant antigens were extracted from inclusion bodies by urea, then refolded and purified by immobilized-metal affinity chromatography (IMAC). Purity was verified by 2D-PAGE and secondary structures were assessed by CD spectroscopy. IgE reactivity of H1sD2 and D2 was tested in western blot with sera from 8 persons with clinical history of HDM allergy. Immunogenicity of H1sD2 and D2 were analyzed in Balb/c mice. Cytokine profile was analyzed by ELISA after stimulation of mouse spleen cells with H1sD2 and D2. Leukocyte population abundance of cells isolated from spleen and lymph node was assessed by flow cytometry. Key findings: Purified recombinant proteins H1sD2 (42 kDa) and D2 (15 kDa) revealed well defined secondary structures, and preserved IgE reactive epitopes. Analysis of supernatants of mouse spleen cells after stimulation with H1sD2 and D2, revealed a qualitatively different cytokine profile from H1sD2 immunized mouse cells (increase in IL10). CD8+ cells were decreased in the lymph node of D2 immunized mice, whereas H1sD2 immunization led to an increase of CD8+ cells in both the lymph node and the spleen. Significance: H1sD2 chimera attenuates Der p 2-inherent Th2 response and directs the immune response toward Th1 and Treg phenotype.
PB  - Oxford : Pergamon-Elsevier Science Ltd
T2  - Life Sciences
T1  - Newly designed hemagglutinin-Der p 2 chimera is a potential candidate for allergen specific immunotherapy
VL  - 213
SP  - 158
EP  - 165
DO  - 10.1016/j.lfs.2018.10.036
ER  - 

@article{
author = "Mrkić, Ivan and Minić, Rajna and Popović, Dragan and Zivkovic, Irena and Gavrović-Jankulović, Marija",
year = "2018",
abstract = "Aim To investigate the immunomodulatory potential of a chimera composed of the receptor-binding domain of hemagglutinin 1 (H1s) from Influenza virus and Der p 2 (D2) allergen for allergen-specific immunotherapy of house-dust mite allergy (HDM). Main methods: H1sD2 chimera and D2 allergen were produced by genetic engineering in E. coli. Recombinant antigens were extracted from inclusion bodies by urea, then refolded and purified by immobilized-metal affinity chromatography (IMAC). Purity was verified by 2D-PAGE and secondary structures were assessed by CD spectroscopy. IgE reactivity of H1sD2 and D2 was tested in western blot with sera from 8 persons with clinical history of HDM allergy. Immunogenicity of H1sD2 and D2 were analyzed in Balb/c mice. Cytokine profile was analyzed by ELISA after stimulation of mouse spleen cells with H1sD2 and D2. Leukocyte population abundance of cells isolated from spleen and lymph node was assessed by flow cytometry. Key findings: Purified recombinant proteins H1sD2 (42 kDa) and D2 (15 kDa) revealed well defined secondary structures, and preserved IgE reactive epitopes. Analysis of supernatants of mouse spleen cells after stimulation with H1sD2 and D2, revealed a qualitatively different cytokine profile from H1sD2 immunized mouse cells (increase in IL10). CD8+ cells were decreased in the lymph node of D2 immunized mice, whereas H1sD2 immunization led to an increase of CD8+ cells in both the lymph node and the spleen. Significance: H1sD2 chimera attenuates Der p 2-inherent Th2 response and directs the immune response toward Th1 and Treg phenotype.",
publisher = "Oxford : Pergamon-Elsevier Science Ltd",
journal = "Life Sciences",
title = "Newly designed hemagglutinin-Der p 2 chimera is a potential candidate for allergen specific immunotherapy",
volume = "213",
pages = "158-165",
doi = "10.1016/j.lfs.2018.10.036"
}

Mrkić, I., Minić, R., Popović, D., Zivkovic, I.,& Gavrović-Jankulović, M.. (2018). Newly designed hemagglutinin-Der p 2 chimera is a potential candidate for allergen specific immunotherapy. in Life Sciences
Oxford : Pergamon-Elsevier Science Ltd., 213, 158-165.
https://doi.org/10.1016/j.lfs.2018.10.036

Mrkić I, Minić R, Popović D, Zivkovic I, Gavrović-Jankulović M. Newly designed hemagglutinin-Der p 2 chimera is a potential candidate for allergen specific immunotherapy. in Life Sciences. 2018;213:158-165.
doi:10.1016/j.lfs.2018.10.036 .

Mrkić, Ivan, Minić, Rajna, Popović, Dragan, Zivkovic, Irena, Gavrović-Jankulović, Marija, "Newly designed hemagglutinin-Der p 2 chimera is a potential candidate for allergen specific immunotherapy" in Life Sciences, 213 (2018):158-165,
https://doi.org/10.1016/j.lfs.2018.10.036 . .

TY  - CONF
AU  - Protić-Rosić, Isidora
AU  - Popović, Milica
AU  - Anđelković, Uroš
AU  - Gavrović-Jankulović, Marija
PY  - 2018
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/3546
AB  - The sublingual route of allergens administration in allergen-specific immunotherapy (ASIT) is proven to be a successful way to treat patients with respiratory allergy. The trend of replacing natural extracts with purified recombinant allergens is growing. Although the purified allergens themselves are not good immunogens, the combined vector systems and adjuvans can improve their immunogenicity 1. Cell surfaces are decorated by different glycan structures, so the lectins specific for these glycans can be used to deliver particular therapeutic to target specific tissue 2. Banana lectin (BanLec) is mannose-specific protein which belongs to the subfamily of Jacalin related lectins 3. Apart from its characteristic to bind glycans, BanLec also modulates immune cells in vitro 4. On the other hand, Bet v 1 (Betula verrucosa) is the major birch pollen allergen. T-cell epitops are distributed over almost entire protein structure 5. In the study the recombinant BanLec-Bet v 1 construct is designed, produced by the recombinant DNA technology, purified and characterized by classical biochemical methods for the application in the ASIT of birch pollen allergy. The expression of newly designed BanLec-Bet v 1 was performed in E. coli BL21 (DE3). After expression the protein was found in the inclusion bodies from which it was extracted with 4 M urea solution. After renaturation, affinity chromatography (Sephadex G-75 superfine) was used for protein purification. Biochemical characterization of the chimera was performed by: SDS PAGE electrophoreses, CD spectroscopy and mass spectrometry. Biological activity of the construct was confirmed by binding of BanLecBet v 1 to a horseradish peroxidase glycoprotein in ELISA. Purified BanLec-Bet v 1 showed molecular mass of 32 kDa. CD spectra of the recombinant construct revealed well defined secondary structures with predominant beta sheets (41.2%). By mass spectrometry 51.8% of the BanLec-Bet v 1 primary structure was confirmed. Biologicaly active recombinant BanLec-Bet v 1 was produced by the recombinant DNA technology. Further in vitro and in vivo studies will evaluate immunomodulatory potential of BanLec-Bet v 1 for application in ASIT.
PB  - Serbian Biochemical Society
C3  - Serbian Biochemical Society Eighth Conference with international participation, “Coordination in Biochemistry and Life”, University of Novi Sad – Rectorate Hall, 16.11.2018. Novi Sad, Serbia
T1  - Production, purification and structural characterisation of recombinant BanLec-Bet v 1
SP  - 177
EP  - 178
UR  - https://hdl.handle.net/21.15107/rcub_cer_3546
ER  - 

@conference{
author = "Protić-Rosić, Isidora and Popović, Milica and Anđelković, Uroš and Gavrović-Jankulović, Marija",
year = "2018",
abstract = "The sublingual route of allergens administration in allergen-specific immunotherapy (ASIT) is proven to be a successful way to treat patients with respiratory allergy. The trend of replacing natural extracts with purified recombinant allergens is growing. Although the purified allergens themselves are not good immunogens, the combined vector systems and adjuvans can improve their immunogenicity 1. Cell surfaces are decorated by different glycan structures, so the lectins specific for these glycans can be used to deliver particular therapeutic to target specific tissue 2. Banana lectin (BanLec) is mannose-specific protein which belongs to the subfamily of Jacalin related lectins 3. Apart from its characteristic to bind glycans, BanLec also modulates immune cells in vitro 4. On the other hand, Bet v 1 (Betula verrucosa) is the major birch pollen allergen. T-cell epitops are distributed over almost entire protein structure 5. In the study the recombinant BanLec-Bet v 1 construct is designed, produced by the recombinant DNA technology, purified and characterized by classical biochemical methods for the application in the ASIT of birch pollen allergy. The expression of newly designed BanLec-Bet v 1 was performed in E. coli BL21 (DE3). After expression the protein was found in the inclusion bodies from which it was extracted with 4 M urea solution. After renaturation, affinity chromatography (Sephadex G-75 superfine) was used for protein purification. Biochemical characterization of the chimera was performed by: SDS PAGE electrophoreses, CD spectroscopy and mass spectrometry. Biological activity of the construct was confirmed by binding of BanLecBet v 1 to a horseradish peroxidase glycoprotein in ELISA. Purified BanLec-Bet v 1 showed molecular mass of 32 kDa. CD spectra of the recombinant construct revealed well defined secondary structures with predominant beta sheets (41.2%). By mass spectrometry 51.8% of the BanLec-Bet v 1 primary structure was confirmed. Biologicaly active recombinant BanLec-Bet v 1 was produced by the recombinant DNA technology. Further in vitro and in vivo studies will evaluate immunomodulatory potential of BanLec-Bet v 1 for application in ASIT.",
publisher = "Serbian Biochemical Society",
journal = "Serbian Biochemical Society Eighth Conference with international participation, “Coordination in Biochemistry and Life”, University of Novi Sad – Rectorate Hall, 16.11.2018. Novi Sad, Serbia",
title = "Production, purification and structural characterisation of recombinant BanLec-Bet v 1",
pages = "177-178",
url = "https://hdl.handle.net/21.15107/rcub_cer_3546"
}

Protić-Rosić, I., Popović, M., Anđelković, U.,& Gavrović-Jankulović, M.. (2018). Production, purification and structural characterisation of recombinant BanLec-Bet v 1. in Serbian Biochemical Society Eighth Conference with international participation, “Coordination in Biochemistry and Life”, University of Novi Sad – Rectorate Hall, 16.11.2018. Novi Sad, Serbia
Serbian Biochemical Society., 177-178.
https://hdl.handle.net/21.15107/rcub_cer_3546

Protić-Rosić I, Popović M, Anđelković U, Gavrović-Jankulović M. Production, purification and structural characterisation of recombinant BanLec-Bet v 1. in Serbian Biochemical Society Eighth Conference with international participation, “Coordination in Biochemistry and Life”, University of Novi Sad – Rectorate Hall, 16.11.2018. Novi Sad, Serbia. 2018;:177-178.
https://hdl.handle.net/21.15107/rcub_cer_3546 .

Protić-Rosić, Isidora, Popović, Milica, Anđelković, Uroš, Gavrović-Jankulović, Marija, "Production, purification and structural characterisation of recombinant BanLec-Bet v 1" in Serbian Biochemical Society Eighth Conference with international participation, “Coordination in Biochemistry and Life”, University of Novi Sad – Rectorate Hall, 16.11.2018. Novi Sad, Serbia (2018):177-178,
https://hdl.handle.net/21.15107/rcub_cer_3546 .

TY  - JOUR
AU  - Mrkić, Ivan
AU  - Minić, Rajna
AU  - Popović, Dragan
AU  - Zivkovic, Irena
AU  - Gavrović-Jankulović, Marija
PY  - 2018
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/4288
AB  - Aim To investigate the immunomodulatory potential of a chimera composed of the receptor-binding domain of hemagglutinin 1 (H1s) from Influenza virus and Der p 2 (D2) allergen for allergen-specific immunotherapy of house-dust mite allergy (HDM). Main methods: H1sD2 chimera and D2 allergen were produced by genetic engineering in E. coli. Recombinant antigens were extracted from inclusion bodies by urea, then refolded and purified by immobilized-metal affinity chromatography (IMAC). Purity was verified by 2D-PAGE and secondary structures were assessed by CD spectroscopy. IgE reactivity of H1sD2 and D2 was tested in western blot with sera from 8 persons with clinical history of HDM allergy. Immunogenicity of H1sD2 and D2 were analyzed in Balb/c mice. Cytokine profile was analyzed by ELISA after stimulation of mouse spleen cells with H1sD2 and D2. Leukocyte population abundance of cells isolated from spleen and lymph node was assessed by flow cytometry. Key findings: Purified recombinant proteins H1sD2 (42 kDa) and D2 (15 kDa) revealed well defined secondary structures, and preserved IgE reactive epitopes. Analysis of supernatants of mouse spleen cells after stimulation with H1sD2 and D2, revealed a qualitatively different cytokine profile from H1sD2 immunized mouse cells (increase in IL10). CD8+ cells were decreased in the lymph node of D2 immunized mice, whereas H1sD2 immunization led to an increase of CD8+ cells in both the lymph node and the spleen. Significance: H1sD2 chimera attenuates Der p 2-inherent Th2 response and directs the immune response toward Th1 and Treg phenotype.
PB  - Oxford : Pergamon-Elsevier Science Ltd
T2  - Life Sciences
T1  - Newly designed hemagglutinin-Der p 2 chimera is a potential candidate for allergen specific immunotherapy
VL  - 213
SP  - 158
EP  - 165
DO  - 10.1016/j.lfs.2018.10.036
ER  - 

@article{
author = "Mrkić, Ivan and Minić, Rajna and Popović, Dragan and Zivkovic, Irena and Gavrović-Jankulović, Marija",
year = "2018",
abstract = "Aim To investigate the immunomodulatory potential of a chimera composed of the receptor-binding domain of hemagglutinin 1 (H1s) from Influenza virus and Der p 2 (D2) allergen for allergen-specific immunotherapy of house-dust mite allergy (HDM). Main methods: H1sD2 chimera and D2 allergen were produced by genetic engineering in E. coli. Recombinant antigens were extracted from inclusion bodies by urea, then refolded and purified by immobilized-metal affinity chromatography (IMAC). Purity was verified by 2D-PAGE and secondary structures were assessed by CD spectroscopy. IgE reactivity of H1sD2 and D2 was tested in western blot with sera from 8 persons with clinical history of HDM allergy. Immunogenicity of H1sD2 and D2 were analyzed in Balb/c mice. Cytokine profile was analyzed by ELISA after stimulation of mouse spleen cells with H1sD2 and D2. Leukocyte population abundance of cells isolated from spleen and lymph node was assessed by flow cytometry. Key findings: Purified recombinant proteins H1sD2 (42 kDa) and D2 (15 kDa) revealed well defined secondary structures, and preserved IgE reactive epitopes. Analysis of supernatants of mouse spleen cells after stimulation with H1sD2 and D2, revealed a qualitatively different cytokine profile from H1sD2 immunized mouse cells (increase in IL10). CD8+ cells were decreased in the lymph node of D2 immunized mice, whereas H1sD2 immunization led to an increase of CD8+ cells in both the lymph node and the spleen. Significance: H1sD2 chimera attenuates Der p 2-inherent Th2 response and directs the immune response toward Th1 and Treg phenotype.",
publisher = "Oxford : Pergamon-Elsevier Science Ltd",
journal = "Life Sciences",
title = "Newly designed hemagglutinin-Der p 2 chimera is a potential candidate for allergen specific immunotherapy",
volume = "213",
pages = "158-165",
doi = "10.1016/j.lfs.2018.10.036"
}

Mrkić, I., Minić, R., Popović, D., Zivkovic, I.,& Gavrović-Jankulović, M.. (2018). Newly designed hemagglutinin-Der p 2 chimera is a potential candidate for allergen specific immunotherapy. in Life Sciences
Oxford : Pergamon-Elsevier Science Ltd., 213, 158-165.
https://doi.org/10.1016/j.lfs.2018.10.036

Mrkić I, Minić R, Popović D, Zivkovic I, Gavrović-Jankulović M. Newly designed hemagglutinin-Der p 2 chimera is a potential candidate for allergen specific immunotherapy. in Life Sciences. 2018;213:158-165.
doi:10.1016/j.lfs.2018.10.036 .

Mrkić, Ivan, Minić, Rajna, Popović, Dragan, Zivkovic, Irena, Gavrović-Jankulović, Marija, "Newly designed hemagglutinin-Der p 2 chimera is a potential candidate for allergen specific immunotherapy" in Life Sciences, 213 (2018):158-165,
https://doi.org/10.1016/j.lfs.2018.10.036 . .

TY  - JOUR
AU  - Nikolić, Jasna
AU  - Nešić, Andrijana
AU  - Čavić, Milena
AU  - Đorđević, Neda O.
AU  - Anđelković, Uroš
AU  - Atanasković-Marković, Marina
AU  - Drakulić, Branko
AU  - Gavrović-Jankulović, Marija
PY  - 2017
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/2058
AB  - Background: Protein oxidation can occur as a consequence of lipid peroxidation during food processing. The aim of this work was to explore the effect of malondialdehyde (MDA) modification of ovalbumin (OVA) on its interaction with T84 intestinal cells. Methods: Molecular dynamics simulation was employed for the prediction of MDA modification in the OVA, while introduced structural changes were evaluated by measurement of carbonyl group content, fluorescence spectra, MS/MS analysis, and IgE reactivity. Effects of MDA modified OVA on T84 epithelial cells were analyzed by gene expression for pro-inflammatory cytokines and protein secretion. Results: Out of 9 predicted, five modified Lys residues were confirmed by MS/MS analysis: (51)TQINKVVR(58), (85)DILNQITKPNDVYSFSLASR(104), (111)YPILPEYLQCVKELYR(126), (187)AFKDEDTQAMPFR(99), (KIKVYLPR284)-K-277, and (IKVYLPR284)-I-278. The introduced MDA modifications influenced profile of IgE reactivity to OVA. Treatment of T84 epithelial cells with OVA and OVA modified with 1 mM MDA, induced up-regulation of pro-inflammatory cytokines (IL-1 beta,IL-25, IL-33, TSLP and TNF alpha), while OVA modification with 10 mM MDA induced down regulation of the cytokine expression profile, except for IL-1 beta. OVA and OVA modified with 1 mM MDA induced secretion of epithelial cells specific cytokine IL-33. Conclusions: This finding indicated that OVA and its MDA modified form have the potential to trigger the innate immunity by inducing up-regulation and secretion of pro-allergenic IL-33 in T84 intestinal epithelial cells. General significance: Interactions of ovalbumin and its MDA modified form with intestinal epithelial cells can induce a specific immunological priming necessary for the downstream activation of innate immunity.
PB  - Elsevier
T2  - Biochimica et Biophysica Acta-General Subjects
T1  - Effect of malondialdehyde on the ovalbumin structure and its interactions with T84 epithelial cells
VL  - 1861
IS  - 2
SP  - 126
EP  - 134
DO  - 10.1016/j.bbagen.2016.11.021
ER  - 

@article{
author = "Nikolić, Jasna and Nešić, Andrijana and Čavić, Milena and Đorđević, Neda O. and Anđelković, Uroš and Atanasković-Marković, Marina and Drakulić, Branko and Gavrović-Jankulović, Marija",
year = "2017",
abstract = "Background: Protein oxidation can occur as a consequence of lipid peroxidation during food processing. The aim of this work was to explore the effect of malondialdehyde (MDA) modification of ovalbumin (OVA) on its interaction with T84 intestinal cells. Methods: Molecular dynamics simulation was employed for the prediction of MDA modification in the OVA, while introduced structural changes were evaluated by measurement of carbonyl group content, fluorescence spectra, MS/MS analysis, and IgE reactivity. Effects of MDA modified OVA on T84 epithelial cells were analyzed by gene expression for pro-inflammatory cytokines and protein secretion. Results: Out of 9 predicted, five modified Lys residues were confirmed by MS/MS analysis: (51)TQINKVVR(58), (85)DILNQITKPNDVYSFSLASR(104), (111)YPILPEYLQCVKELYR(126), (187)AFKDEDTQAMPFR(99), (KIKVYLPR284)-K-277, and (IKVYLPR284)-I-278. The introduced MDA modifications influenced profile of IgE reactivity to OVA. Treatment of T84 epithelial cells with OVA and OVA modified with 1 mM MDA, induced up-regulation of pro-inflammatory cytokines (IL-1 beta,IL-25, IL-33, TSLP and TNF alpha), while OVA modification with 10 mM MDA induced down regulation of the cytokine expression profile, except for IL-1 beta. OVA and OVA modified with 1 mM MDA induced secretion of epithelial cells specific cytokine IL-33. Conclusions: This finding indicated that OVA and its MDA modified form have the potential to trigger the innate immunity by inducing up-regulation and secretion of pro-allergenic IL-33 in T84 intestinal epithelial cells. General significance: Interactions of ovalbumin and its MDA modified form with intestinal epithelial cells can induce a specific immunological priming necessary for the downstream activation of innate immunity.",
publisher = "Elsevier",
journal = "Biochimica et Biophysica Acta-General Subjects",
title = "Effect of malondialdehyde on the ovalbumin structure and its interactions with T84 epithelial cells",
volume = "1861",
number = "2",
pages = "126-134",
doi = "10.1016/j.bbagen.2016.11.021"
}

Nikolić, J., Nešić, A., Čavić, M., Đorđević, N. O., Anđelković, U., Atanasković-Marković, M., Drakulić, B.,& Gavrović-Jankulović, M.. (2017). Effect of malondialdehyde on the ovalbumin structure and its interactions with T84 epithelial cells. in Biochimica et Biophysica Acta-General Subjects
Elsevier., 1861(2), 126-134.
https://doi.org/10.1016/j.bbagen.2016.11.021

Nikolić J, Nešić A, Čavić M, Đorđević NO, Anđelković U, Atanasković-Marković M, Drakulić B, Gavrović-Jankulović M. Effect of malondialdehyde on the ovalbumin structure and its interactions with T84 epithelial cells. in Biochimica et Biophysica Acta-General Subjects. 2017;1861(2):126-134.
doi:10.1016/j.bbagen.2016.11.021 .

Nikolić, Jasna, Nešić, Andrijana, Čavić, Milena, Đorđević, Neda O., Anđelković, Uroš, Atanasković-Marković, Marina, Drakulić, Branko, Gavrović-Jankulović, Marija, "Effect of malondialdehyde on the ovalbumin structure and its interactions with T84 epithelial cells" in Biochimica et Biophysica Acta-General Subjects, 1861, no. 2 (2017):126-134,
https://doi.org/10.1016/j.bbagen.2016.11.021 . .

TY  - JOUR
AU  - Anđelković, Uroš
AU  - Gavrović-Jankulović, Marija
AU  - Martinović, Tamara
AU  - Josić, Djuro
PY  - 2017
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/2113
AB  - Use of foodomics, mostly proteomic and genomic based methods, for study of allergens in food is presented. Immunological methods and nucleic acid-based methods are still most frequently used for diagnosis of allergies and for qualitative and quantitative determination of food allergens. They are sensitive, and can be used for the determination of allergens in trace concentrations. However, lack of specificity and cross-reaction of some antibodies can still be a relevant source of bias. The epitopes of protein allergens with posttranslational modifications and their changes originated during food processing cannot be traced by use of nucleic acid-based strategies. Recent developments of both antibody and nucleic acid-based biosensors, their miniaturization and increasing application of nanotechnology, significantly supported further use of both strategies. Regarding accuracy, reliability and sensitivity, mass spectrometry-based methods bring important advantage over both above presented strategies. Furthermore, the increasing use of mass spectrometry (MS) is discussed. Combined with proper sample preparation, liquid chromatography (LC) and/or different electrophoretic methods, targeted approach in mass spectrometry-based allergen analysis brings an additional strategic advance. However, MS is still rarely used for high-throughput analyses and detection and quantification of allergens for the reasons of price and relatively long time necessary for analysis. Recent developments of new high-resolution instruments are encouraging and enable development in the direction of a high-throughput strategy. Consequently, fast, very sensitive, reliable and accurate detection and quantification of allergens in highly complex samples such as food matrices, and the use of MS in routine determination of allergens can be reached in near future.
PB  - Elsevier Sci Ltd, Oxford
T2  - Trac-Trends in Analytical Chemistry
T1  - Omics methods as a tool for investigation of food allergies
VL  - 96
SP  - 107
EP  - 115
DO  - 10.1016/j.trac.2017.07.011
ER  - 

@article{
author = "Anđelković, Uroš and Gavrović-Jankulović, Marija and Martinović, Tamara and Josić, Djuro",
year = "2017",
abstract = "Use of foodomics, mostly proteomic and genomic based methods, for study of allergens in food is presented. Immunological methods and nucleic acid-based methods are still most frequently used for diagnosis of allergies and for qualitative and quantitative determination of food allergens. They are sensitive, and can be used for the determination of allergens in trace concentrations. However, lack of specificity and cross-reaction of some antibodies can still be a relevant source of bias. The epitopes of protein allergens with posttranslational modifications and their changes originated during food processing cannot be traced by use of nucleic acid-based strategies. Recent developments of both antibody and nucleic acid-based biosensors, their miniaturization and increasing application of nanotechnology, significantly supported further use of both strategies. Regarding accuracy, reliability and sensitivity, mass spectrometry-based methods bring important advantage over both above presented strategies. Furthermore, the increasing use of mass spectrometry (MS) is discussed. Combined with proper sample preparation, liquid chromatography (LC) and/or different electrophoretic methods, targeted approach in mass spectrometry-based allergen analysis brings an additional strategic advance. However, MS is still rarely used for high-throughput analyses and detection and quantification of allergens for the reasons of price and relatively long time necessary for analysis. Recent developments of new high-resolution instruments are encouraging and enable development in the direction of a high-throughput strategy. Consequently, fast, very sensitive, reliable and accurate detection and quantification of allergens in highly complex samples such as food matrices, and the use of MS in routine determination of allergens can be reached in near future.",
publisher = "Elsevier Sci Ltd, Oxford",
journal = "Trac-Trends in Analytical Chemistry",
title = "Omics methods as a tool for investigation of food allergies",
volume = "96",
pages = "107-115",
doi = "10.1016/j.trac.2017.07.011"
}

Anđelković, U., Gavrović-Jankulović, M., Martinović, T.,& Josić, D.. (2017). Omics methods as a tool for investigation of food allergies. in Trac-Trends in Analytical Chemistry
Elsevier Sci Ltd, Oxford., 96, 107-115.
https://doi.org/10.1016/j.trac.2017.07.011

Anđelković U, Gavrović-Jankulović M, Martinović T, Josić D. Omics methods as a tool for investigation of food allergies. in Trac-Trends in Analytical Chemistry. 2017;96:107-115.
doi:10.1016/j.trac.2017.07.011 .

Anđelković, Uroš, Gavrović-Jankulović, Marija, Martinović, Tamara, Josić, Djuro, "Omics methods as a tool for investigation of food allergies" in Trac-Trends in Analytical Chemistry, 96 (2017):107-115,
https://doi.org/10.1016/j.trac.2017.07.011 . .

TY  - JOUR
AU  - Anđelković, Uroš
AU  - Gavrović-Jankulović, Marija
AU  - Martinović, Tamara
AU  - Josić, Djuro
PY  - 2017
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/2937
AB  - Use of foodomics, mostly proteomic and genomic based methods, for study of allergens in food is presented. Immunological methods and nucleic acid-based methods are still most frequently used for diagnosis of allergies and for qualitative and quantitative determination of food allergens. They are sensitive, and can be used for the determination of allergens in trace concentrations. However, lack of specificity and cross-reaction of some antibodies can still be a relevant source of bias. The epitopes of protein allergens with posttranslational modifications and their changes originated during food processing cannot be traced by use of nucleic acid-based strategies. Recent developments of both antibody and nucleic acid-based biosensors, their miniaturization and increasing application of nanotechnology, significantly supported further use of both strategies. Regarding accuracy, reliability and sensitivity, mass spectrometry-based methods bring important advantage over both above presented strategies. Furthermore, the increasing use of mass spectrometry (MS) is discussed. Combined with proper sample preparation, liquid chromatography (LC) and/or different electrophoretic methods, targeted approach in mass spectrometry-based allergen analysis brings an additional strategic advance. However, MS is still rarely used for high-throughput analyses and detection and quantification of allergens for the reasons of price and relatively long time necessary for analysis. Recent developments of new high-resolution instruments are encouraging and enable development in the direction of a high-throughput strategy. Consequently, fast, very sensitive, reliable and accurate detection and quantification of allergens in highly complex samples such as food matrices, and the use of MS in routine determination of allergens can be reached in near future.
PB  - Elsevier
T2  - Trac-Trends in Analytical Chemistry
T1  - Omics methods as a tool for investigation of food allergies
VL  - 96
SP  - 107
EP  - 115
DO  - 10.1016/j.trac.2017.07.011
ER  - 

@article{
author = "Anđelković, Uroš and Gavrović-Jankulović, Marija and Martinović, Tamara and Josić, Djuro",
year = "2017",
abstract = "Use of foodomics, mostly proteomic and genomic based methods, for study of allergens in food is presented. Immunological methods and nucleic acid-based methods are still most frequently used for diagnosis of allergies and for qualitative and quantitative determination of food allergens. They are sensitive, and can be used for the determination of allergens in trace concentrations. However, lack of specificity and cross-reaction of some antibodies can still be a relevant source of bias. The epitopes of protein allergens with posttranslational modifications and their changes originated during food processing cannot be traced by use of nucleic acid-based strategies. Recent developments of both antibody and nucleic acid-based biosensors, their miniaturization and increasing application of nanotechnology, significantly supported further use of both strategies. Regarding accuracy, reliability and sensitivity, mass spectrometry-based methods bring important advantage over both above presented strategies. Furthermore, the increasing use of mass spectrometry (MS) is discussed. Combined with proper sample preparation, liquid chromatography (LC) and/or different electrophoretic methods, targeted approach in mass spectrometry-based allergen analysis brings an additional strategic advance. However, MS is still rarely used for high-throughput analyses and detection and quantification of allergens for the reasons of price and relatively long time necessary for analysis. Recent developments of new high-resolution instruments are encouraging and enable development in the direction of a high-throughput strategy. Consequently, fast, very sensitive, reliable and accurate detection and quantification of allergens in highly complex samples such as food matrices, and the use of MS in routine determination of allergens can be reached in near future.",
publisher = "Elsevier",
journal = "Trac-Trends in Analytical Chemistry",
title = "Omics methods as a tool for investigation of food allergies",
volume = "96",
pages = "107-115",
doi = "10.1016/j.trac.2017.07.011"
}

Anđelković, U., Gavrović-Jankulović, M., Martinović, T.,& Josić, D.. (2017). Omics methods as a tool for investigation of food allergies. in Trac-Trends in Analytical Chemistry
Elsevier., 96, 107-115.
https://doi.org/10.1016/j.trac.2017.07.011

Anđelković U, Gavrović-Jankulović M, Martinović T, Josić D. Omics methods as a tool for investigation of food allergies. in Trac-Trends in Analytical Chemistry. 2017;96:107-115.
doi:10.1016/j.trac.2017.07.011 .

Anđelković, Uroš, Gavrović-Jankulović, Marija, Martinović, Tamara, Josić, Djuro, "Omics methods as a tool for investigation of food allergies" in Trac-Trends in Analytical Chemistry, 96 (2017):107-115,
https://doi.org/10.1016/j.trac.2017.07.011 . .

TY  - JOUR
AU  - Nikolić, Jasna
AU  - Nešić, Andrijana
AU  - Čavić, Milena
AU  - Đorđević, Neda O.
AU  - Anđelković, Uroš
AU  - Atanasković-Marković, Marina
AU  - Drakulić, Branko
AU  - Gavrović-Jankulović, Marija
PY  - 2017
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/3042
AB  - Background: Protein oxidation can occur as a consequence of lipid peroxidation during food processing. The aim of this work was to explore the effect of malondialdehyde (MDA) modification of ovalbumin (OVA) on its interaction with T84 intestinal cells. Methods: Molecular dynamics simulation was employed for the prediction of MDA modification in the OVA, while introduced structural changes were evaluated by measurement of carbonyl group content, fluorescence spectra, MS/MS analysis, and IgE reactivity. Effects of MDA modified OVA on T84 epithelial cells were analyzed by gene expression for pro-inflammatory cytokines and protein secretion. Results: Out of 9 predicted, five modified Lys residues were confirmed by MS/MS analysis: (51)TQINKVVR(58), (85)DILNQITKPNDVYSFSLASR(104), (111)YPILPEYLQCVKELYR(126), (187)AFKDEDTQAMPFR(99), (KIKVYLPR284)-K-277, and (IKVYLPR284)-I-278. The introduced MDA modifications influenced profile of IgE reactivity to OVA. Treatment of T84 epithelial cells with OVA and OVA modified with 1 mM MDA, induced up-regulation of pro-inflammatory cytokines (IL-1 beta,IL-25, IL-33, TSLP and TNF alpha), while OVA modification with 10 mM MDA induced down regulation of the cytokine expression profile, except for IL-1 beta. OVA and OVA modified with 1 mM MDA induced secretion of epithelial cells specific cytokine IL-33. Conclusions: This finding indicated that OVA and its MDA modified form have the potential to trigger the innate immunity by inducing up-regulation and secretion of pro-allergenic IL-33 in T84 intestinal epithelial cells. General significance: Interactions of ovalbumin and its MDA modified form with intestinal epithelial cells can induce a specific immunological priming necessary for the downstream activation of innate immunity.
PB  - Elsevier
T2  - Biochimica et Biophysica Acta-General Subjects
T1  - Effect of malondialdehyde on the ovalbumin structure and its interactions with T84 epithelial cells
VL  - 1861
IS  - 2
SP  - 126
EP  - 134
DO  - 10.1016/j.bbagen.2016.11.021
ER  - 

@article{
author = "Nikolić, Jasna and Nešić, Andrijana and Čavić, Milena and Đorđević, Neda O. and Anđelković, Uroš and Atanasković-Marković, Marina and Drakulić, Branko and Gavrović-Jankulović, Marija",
year = "2017",
abstract = "Background: Protein oxidation can occur as a consequence of lipid peroxidation during food processing. The aim of this work was to explore the effect of malondialdehyde (MDA) modification of ovalbumin (OVA) on its interaction with T84 intestinal cells. Methods: Molecular dynamics simulation was employed for the prediction of MDA modification in the OVA, while introduced structural changes were evaluated by measurement of carbonyl group content, fluorescence spectra, MS/MS analysis, and IgE reactivity. Effects of MDA modified OVA on T84 epithelial cells were analyzed by gene expression for pro-inflammatory cytokines and protein secretion. Results: Out of 9 predicted, five modified Lys residues were confirmed by MS/MS analysis: (51)TQINKVVR(58), (85)DILNQITKPNDVYSFSLASR(104), (111)YPILPEYLQCVKELYR(126), (187)AFKDEDTQAMPFR(99), (KIKVYLPR284)-K-277, and (IKVYLPR284)-I-278. The introduced MDA modifications influenced profile of IgE reactivity to OVA. Treatment of T84 epithelial cells with OVA and OVA modified with 1 mM MDA, induced up-regulation of pro-inflammatory cytokines (IL-1 beta,IL-25, IL-33, TSLP and TNF alpha), while OVA modification with 10 mM MDA induced down regulation of the cytokine expression profile, except for IL-1 beta. OVA and OVA modified with 1 mM MDA induced secretion of epithelial cells specific cytokine IL-33. Conclusions: This finding indicated that OVA and its MDA modified form have the potential to trigger the innate immunity by inducing up-regulation and secretion of pro-allergenic IL-33 in T84 intestinal epithelial cells. General significance: Interactions of ovalbumin and its MDA modified form with intestinal epithelial cells can induce a specific immunological priming necessary for the downstream activation of innate immunity.",
publisher = "Elsevier",
journal = "Biochimica et Biophysica Acta-General Subjects",
title = "Effect of malondialdehyde on the ovalbumin structure and its interactions with T84 epithelial cells",
volume = "1861",
number = "2",
pages = "126-134",
doi = "10.1016/j.bbagen.2016.11.021"
}

Nikolić, J., Nešić, A., Čavić, M., Đorđević, N. O., Anđelković, U., Atanasković-Marković, M., Drakulić, B.,& Gavrović-Jankulović, M.. (2017). Effect of malondialdehyde on the ovalbumin structure and its interactions with T84 epithelial cells. in Biochimica et Biophysica Acta-General Subjects
Elsevier., 1861(2), 126-134.
https://doi.org/10.1016/j.bbagen.2016.11.021

Nikolić J, Nešić A, Čavić M, Đorđević NO, Anđelković U, Atanasković-Marković M, Drakulić B, Gavrović-Jankulović M. Effect of malondialdehyde on the ovalbumin structure and its interactions with T84 epithelial cells. in Biochimica et Biophysica Acta-General Subjects. 2017;1861(2):126-134.
doi:10.1016/j.bbagen.2016.11.021 .

Nikolić, Jasna, Nešić, Andrijana, Čavić, Milena, Đorđević, Neda O., Anđelković, Uroš, Atanasković-Marković, Marina, Drakulić, Branko, Gavrović-Jankulović, Marija, "Effect of malondialdehyde on the ovalbumin structure and its interactions with T84 epithelial cells" in Biochimica et Biophysica Acta-General Subjects, 1861, no. 2 (2017):126-134,
https://doi.org/10.1016/j.bbagen.2016.11.021 . .

TY  - JOUR
AU  - Anđelković, Uroš
AU  - Tufegdžić, Srđan
AU  - Popović, Milica
PY  - 2017
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/2122
AB  - The exclusive properties of monolithic supports enable fast mass transfer, high porosity, low back pressure, easy preparation process and miniaturisation, and the availability of different chemistries make them particularly suitable materials for high-throughput (HTP) protein and peptide separation. In this review recent advances in monolith-based chromatographic supports for HTP screening of protein and peptide samples are presented and their application in HTP sample preparation (separation, enrichment, depletion, proteolytic digestion) for HTP proteomics is discussed. Development and applications of different monolithic capillary columns in HTP MS-based bottom-up and top-down proteomics are overviewed. By discussing the chromatographic conditions and the mass spectrometric data acquisition conditions an attempt is made to present currently demonstrated capacities of monolithic capillary columns for HTP identification and quantification of proteins and peptides from complex biological samples by MS-based proteomics. Some recent advances in basic monolith technology of importance for proteomics are also discussed.
PB  - Wiley, Hoboken
T2  - Electrophoresis
T1  - Use of monolithic supports for high-throughput protein and peptide separation in proteomics
VL  - 38
IS  - 22-23
SP  - 2851
EP  - 2869
DO  - 10.1002/elps.201700260
ER  - 

@article{
author = "Anđelković, Uroš and Tufegdžić, Srđan and Popović, Milica",
year = "2017",
abstract = "The exclusive properties of monolithic supports enable fast mass transfer, high porosity, low back pressure, easy preparation process and miniaturisation, and the availability of different chemistries make them particularly suitable materials for high-throughput (HTP) protein and peptide separation. In this review recent advances in monolith-based chromatographic supports for HTP screening of protein and peptide samples are presented and their application in HTP sample preparation (separation, enrichment, depletion, proteolytic digestion) for HTP proteomics is discussed. Development and applications of different monolithic capillary columns in HTP MS-based bottom-up and top-down proteomics are overviewed. By discussing the chromatographic conditions and the mass spectrometric data acquisition conditions an attempt is made to present currently demonstrated capacities of monolithic capillary columns for HTP identification and quantification of proteins and peptides from complex biological samples by MS-based proteomics. Some recent advances in basic monolith technology of importance for proteomics are also discussed.",
publisher = "Wiley, Hoboken",
journal = "Electrophoresis",
title = "Use of monolithic supports for high-throughput protein and peptide separation in proteomics",
volume = "38",
number = "22-23",
pages = "2851-2869",
doi = "10.1002/elps.201700260"
}

Anđelković, U., Tufegdžić, S.,& Popović, M.. (2017). Use of monolithic supports for high-throughput protein and peptide separation in proteomics. in Electrophoresis
Wiley, Hoboken., 38(22-23), 2851-2869.
https://doi.org/10.1002/elps.201700260

Anđelković U, Tufegdžić S, Popović M. Use of monolithic supports for high-throughput protein and peptide separation in proteomics. in Electrophoresis. 2017;38(22-23):2851-2869.
doi:10.1002/elps.201700260 .

Anđelković, Uroš, Tufegdžić, Srđan, Popović, Milica, "Use of monolithic supports for high-throughput protein and peptide separation in proteomics" in Electrophoresis, 38, no. 22-23 (2017):2851-2869,
https://doi.org/10.1002/elps.201700260 . .

TY  - JOUR
AU  - Anđelković, Uroš
AU  - Tufegdžić, Srđan
AU  - Popović, Milica
PY  - 2017
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/2985
AB  - The exclusive properties of monolithic supports enable fast mass transfer, high porosity, low back pressure, easy preparation process and miniaturisation, and the availability of different chemistries make them particularly suitable materials for high-throughput (HTP) protein and peptide separation. In this review recent advances in monolith-based chromatographic supports for HTP screening of protein and peptide samples are presented and their application in HTP sample preparation (separation, enrichment, depletion, proteolytic digestion) for HTP proteomics is discussed. Development and applications of different monolithic capillary columns in HTP MS-based bottom-up and top-down proteomics are overviewed. By discussing the chromatographic conditions and the mass spectrometric data acquisition conditions an attempt is made to present currently demonstrated capacities of monolithic capillary columns for HTP identification and quantification of proteins and peptides from complex biological samples by MS-based proteomics. Some recent advances in basic monolith technology of importance for proteomics are also discussed.
PB  - Wiley, Hoboken
T2  - Electrophoresis
T1  - Use of monolithic supports for high-throughput protein and peptide separation in proteomics
VL  - 38
IS  - 22-23
SP  - 2851
EP  - 2869
DO  - 10.1002/elps.201700260
ER  - 

@article{
author = "Anđelković, Uroš and Tufegdžić, Srđan and Popović, Milica",
year = "2017",
abstract = "The exclusive properties of monolithic supports enable fast mass transfer, high porosity, low back pressure, easy preparation process and miniaturisation, and the availability of different chemistries make them particularly suitable materials for high-throughput (HTP) protein and peptide separation. In this review recent advances in monolith-based chromatographic supports for HTP screening of protein and peptide samples are presented and their application in HTP sample preparation (separation, enrichment, depletion, proteolytic digestion) for HTP proteomics is discussed. Development and applications of different monolithic capillary columns in HTP MS-based bottom-up and top-down proteomics are overviewed. By discussing the chromatographic conditions and the mass spectrometric data acquisition conditions an attempt is made to present currently demonstrated capacities of monolithic capillary columns for HTP identification and quantification of proteins and peptides from complex biological samples by MS-based proteomics. Some recent advances in basic monolith technology of importance for proteomics are also discussed.",
publisher = "Wiley, Hoboken",
journal = "Electrophoresis",
title = "Use of monolithic supports for high-throughput protein and peptide separation in proteomics",
volume = "38",
number = "22-23",
pages = "2851-2869",
doi = "10.1002/elps.201700260"
}

Anđelković, U., Tufegdžić, S.,& Popović, M.. (2017). Use of monolithic supports for high-throughput protein and peptide separation in proteomics. in Electrophoresis
Wiley, Hoboken., 38(22-23), 2851-2869.
https://doi.org/10.1002/elps.201700260

Anđelković U, Tufegdžić S, Popović M. Use of monolithic supports for high-throughput protein and peptide separation in proteomics. in Electrophoresis. 2017;38(22-23):2851-2869.
doi:10.1002/elps.201700260 .

Anđelković, Uroš, Tufegdžić, Srđan, Popović, Milica, "Use of monolithic supports for high-throughput protein and peptide separation in proteomics" in Electrophoresis, 38, no. 22-23 (2017):2851-2869,
https://doi.org/10.1002/elps.201700260 . .

TY  - JOUR
AU  - Grozdanović, Milica
AU  - Čavić, Milena
AU  - Nešić, Andrijana
AU  - Anđelković, Uroš
AU  - Akbari, Peyman
AU  - Smit, Joost J.
AU  - Gavrović-Jankulović, Marija
PY  - 2016
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/3154
AB  - Background: The intestinal epithelium forms a barrier that food allergens must cross in order to induce sensitization. The aim of this study was to evaluate the impact of the plant-derived food cysteine protease - actinidin (Act d1) on the integrity of intestinal epithelium tight junctions (TJs). Methods: Effects of Act d1 on the intestinal epithelium were evaluated in Caco-2 monolayers and in a mouse model by measuring transepithelial resistance and in vivo permeability. Integrity of the tight junctions was analyzed by confocal microscopy. Proteolysis of TJ protein occludin was evaluated by mass spectrometry. Results: Actinidin (1 mg/mL) reduced the transepithelial resistance of the cell monolayer by 18.1% (after 1 h) and 25.6% (after 4 h). This loss of barrier function was associated with Act d 1 disruption of the occludin and zonula occludens (ZO)-1 network. The effect on intestinal permeability in vivo was demonstrated by the significantly higher concentration of 40 kDa FITC-dextran (233 mu g/mL) that passed from the intestine into the serum of Act d1 treated mice in comparison to the control group (0.5 mu g/mL). Human occludin was fragmented, and putative Act d1 cleavage sites were identified in extracellular loops of human occludin. Conclusion: Act d1 caused protease-dependent disruption of tight junctions in confluent Caco-2 cells and increased intestinal permeability in mice. General significance: In line with the observed effects of food cysteine proteases in occupational allergy, these results suggest that disruption of tight junctions by food cysteine proteases may contribute to the process of sensitization in food allergy. (C) 2015 Elsevier B.V. All rights reserved.
PB  - Elsevier
T2  - Biochimica et Biophysica Acta: General Subjects
T1  - Kiwifruit cysteine protease actinidin compromises the intestinal barrier by disrupting tight junctions
VL  - 1860
IS  - 3
SP  - 516
EP  - 526
DO  - 10.1016/j.bbagen.2015.12.005
ER  - 

@article{
author = "Grozdanović, Milica and Čavić, Milena and Nešić, Andrijana and Anđelković, Uroš and Akbari, Peyman and Smit, Joost J. and Gavrović-Jankulović, Marija",
year = "2016",
abstract = "Background: The intestinal epithelium forms a barrier that food allergens must cross in order to induce sensitization. The aim of this study was to evaluate the impact of the plant-derived food cysteine protease - actinidin (Act d1) on the integrity of intestinal epithelium tight junctions (TJs). Methods: Effects of Act d1 on the intestinal epithelium were evaluated in Caco-2 monolayers and in a mouse model by measuring transepithelial resistance and in vivo permeability. Integrity of the tight junctions was analyzed by confocal microscopy. Proteolysis of TJ protein occludin was evaluated by mass spectrometry. Results: Actinidin (1 mg/mL) reduced the transepithelial resistance of the cell monolayer by 18.1% (after 1 h) and 25.6% (after 4 h). This loss of barrier function was associated with Act d 1 disruption of the occludin and zonula occludens (ZO)-1 network. The effect on intestinal permeability in vivo was demonstrated by the significantly higher concentration of 40 kDa FITC-dextran (233 mu g/mL) that passed from the intestine into the serum of Act d1 treated mice in comparison to the control group (0.5 mu g/mL). Human occludin was fragmented, and putative Act d1 cleavage sites were identified in extracellular loops of human occludin. Conclusion: Act d1 caused protease-dependent disruption of tight junctions in confluent Caco-2 cells and increased intestinal permeability in mice. General significance: In line with the observed effects of food cysteine proteases in occupational allergy, these results suggest that disruption of tight junctions by food cysteine proteases may contribute to the process of sensitization in food allergy. (C) 2015 Elsevier B.V. All rights reserved.",
publisher = "Elsevier",
journal = "Biochimica et Biophysica Acta: General Subjects",
title = "Kiwifruit cysteine protease actinidin compromises the intestinal barrier by disrupting tight junctions",
volume = "1860",
number = "3",
pages = "516-526",
doi = "10.1016/j.bbagen.2015.12.005"
}

Grozdanović, M., Čavić, M., Nešić, A., Anđelković, U., Akbari, P., Smit, J. J.,& Gavrović-Jankulović, M.. (2016). Kiwifruit cysteine protease actinidin compromises the intestinal barrier by disrupting tight junctions. in Biochimica et Biophysica Acta: General Subjects
Elsevier., 1860(3), 516-526.
https://doi.org/10.1016/j.bbagen.2015.12.005

Grozdanović M, Čavić M, Nešić A, Anđelković U, Akbari P, Smit JJ, Gavrović-Jankulović M. Kiwifruit cysteine protease actinidin compromises the intestinal barrier by disrupting tight junctions. in Biochimica et Biophysica Acta: General Subjects. 2016;1860(3):516-526.
doi:10.1016/j.bbagen.2015.12.005 .

Grozdanović, Milica, Čavić, Milena, Nešić, Andrijana, Anđelković, Uroš, Akbari, Peyman, Smit, Joost J., Gavrović-Jankulović, Marija, "Kiwifruit cysteine protease actinidin compromises the intestinal barrier by disrupting tight junctions" in Biochimica et Biophysica Acta: General Subjects, 1860, no. 3 (2016):516-526,
https://doi.org/10.1016/j.bbagen.2015.12.005 . .

TY  - JOUR
AU  - Grozdanović, Milica
AU  - Čavić, Milena
AU  - Nešić, Andrijana
AU  - Anđelković, Uroš
AU  - Akbari, Peyman
AU  - Smit, Joost J.
AU  - Gavrović-Jankulović, Marija
PY  - 2016
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/3155
AB  - Background: The intestinal epithelium forms a barrier that food allergens must cross in order to induce sensitization. The aim of this study was to evaluate the impact of the plant-derived food cysteine protease - actinidin (Act d1) on the integrity of intestinal epithelium tight junctions (TJs). Methods: Effects of Act d1 on the intestinal epithelium were evaluated in Caco-2 monolayers and in a mouse model by measuring transepithelial resistance and in vivo permeability. Integrity of the tight junctions was analyzed by confocal microscopy. Proteolysis of TJ protein occludin was evaluated by mass spectrometry. Results: Actinidin (1 mg/mL) reduced the transepithelial resistance of the cell monolayer by 18.1% (after 1 h) and 25.6% (after 4 h). This loss of barrier function was associated with Act d 1 disruption of the occludin and zonula occludens (ZO)-1 network. The effect on intestinal permeability in vivo was demonstrated by the significantly higher concentration of 40 kDa FITC-dextran (233 mu g/mL) that passed from the intestine into the serum of Act d1 treated mice in comparison to the control group (0.5 mu g/mL). Human occludin was fragmented, and putative Act d1 cleavage sites were identified in extracellular loops of human occludin. Conclusion: Act d1 caused protease-dependent disruption of tight junctions in confluent Caco-2 cells and increased intestinal permeability in mice. General significance: In line with the observed effects of food cysteine proteases in occupational allergy, these results suggest that disruption of tight junctions by food cysteine proteases may contribute to the process of sensitization in food allergy. (C) 2015 Elsevier B.V. All rights reserved.
PB  - Elsevier
T2  - Biochimica et Biophysica Acta: General Subjects
T1  - Kiwifruit cysteine protease actinidin compromises the intestinal barrier by disrupting tight junctions
VL  - 1860
IS  - 3
SP  - 516
EP  - 526
DO  - 10.1016/j.bbagen.2015.12.005
ER  - 

@article{
author = "Grozdanović, Milica and Čavić, Milena and Nešić, Andrijana and Anđelković, Uroš and Akbari, Peyman and Smit, Joost J. and Gavrović-Jankulović, Marija",
year = "2016",
abstract = "Background: The intestinal epithelium forms a barrier that food allergens must cross in order to induce sensitization. The aim of this study was to evaluate the impact of the plant-derived food cysteine protease - actinidin (Act d1) on the integrity of intestinal epithelium tight junctions (TJs). Methods: Effects of Act d1 on the intestinal epithelium were evaluated in Caco-2 monolayers and in a mouse model by measuring transepithelial resistance and in vivo permeability. Integrity of the tight junctions was analyzed by confocal microscopy. Proteolysis of TJ protein occludin was evaluated by mass spectrometry. Results: Actinidin (1 mg/mL) reduced the transepithelial resistance of the cell monolayer by 18.1% (after 1 h) and 25.6% (after 4 h). This loss of barrier function was associated with Act d 1 disruption of the occludin and zonula occludens (ZO)-1 network. The effect on intestinal permeability in vivo was demonstrated by the significantly higher concentration of 40 kDa FITC-dextran (233 mu g/mL) that passed from the intestine into the serum of Act d1 treated mice in comparison to the control group (0.5 mu g/mL). Human occludin was fragmented, and putative Act d1 cleavage sites were identified in extracellular loops of human occludin. Conclusion: Act d1 caused protease-dependent disruption of tight junctions in confluent Caco-2 cells and increased intestinal permeability in mice. General significance: In line with the observed effects of food cysteine proteases in occupational allergy, these results suggest that disruption of tight junctions by food cysteine proteases may contribute to the process of sensitization in food allergy. (C) 2015 Elsevier B.V. All rights reserved.",
publisher = "Elsevier",
journal = "Biochimica et Biophysica Acta: General Subjects",
title = "Kiwifruit cysteine protease actinidin compromises the intestinal barrier by disrupting tight junctions",
volume = "1860",
number = "3",
pages = "516-526",
doi = "10.1016/j.bbagen.2015.12.005"
}

Grozdanović, M., Čavić, M., Nešić, A., Anđelković, U., Akbari, P., Smit, J. J.,& Gavrović-Jankulović, M.. (2016). Kiwifruit cysteine protease actinidin compromises the intestinal barrier by disrupting tight junctions. in Biochimica et Biophysica Acta: General Subjects
Elsevier., 1860(3), 516-526.
https://doi.org/10.1016/j.bbagen.2015.12.005

Grozdanović M, Čavić M, Nešić A, Anđelković U, Akbari P, Smit JJ, Gavrović-Jankulović M. Kiwifruit cysteine protease actinidin compromises the intestinal barrier by disrupting tight junctions. in Biochimica et Biophysica Acta: General Subjects. 2016;1860(3):516-526.
doi:10.1016/j.bbagen.2015.12.005 .

Grozdanović, Milica, Čavić, Milena, Nešić, Andrijana, Anđelković, Uroš, Akbari, Peyman, Smit, Joost J., Gavrović-Jankulović, Marija, "Kiwifruit cysteine protease actinidin compromises the intestinal barrier by disrupting tight junctions" in Biochimica et Biophysica Acta: General Subjects, 1860, no. 3 (2016):516-526,
https://doi.org/10.1016/j.bbagen.2015.12.005 . .

TY  - JOUR
AU  - Prodanović, Olivera
AU  - Spasojevic, Dragica
AU  - Prokopijevic, Milos
AU  - Radotić, Ksenija
AU  - Marković, Nevena
AU  - Blažić, Marija
AU  - Prodanović, Radivoje
PY  - 2015
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/1727
AB  - Phenol and amino groups were introduced into alginate to different degrees via oxidation with 2.5, 5, 10, 15 and 20 mol% of periodate and reductive amination by tyramine. Modification of alginate with tyramine was confirmed by FTIR spectroscopy and UV-VIS spectroscopy, while concentration of phenol and ionizable groups was determined using absorbance at 275 nm and acid-base titration. All tyramine-alginates were able to form hydrogels after cross-linking with horse radish peroxidase (HRP) and hydrogen peroxide. Tyramine-alginates oxidized with up to 10 mol% of periodate were also capable of forming hydrogels with calcium ions. Tyramine-alginates were tested for HRP immobilization within micro-beads obtained by peroxidase catalyzed droplet polymerization using internal delivery of hydrogen peroxide via glucose oxidase and glucose. Highest activity of immobilized peroxidase was obtained with 20% (w/v) tyramine-alginate obtained via 20 mol% periodate oxidation. Immobilized enzyme was not leaking from the micro-beads and was further kinetically characterized for pyrogallol oxidation. Km for pyrogallol was increased after immobilization from 1.93 mM for soluble HRP to 734 mM for immobilized HRP. The optimum pH was also increased from pH 7.0 to 8.0. Temperature and organic solvent stability improved significantly after immobilization, so that half-life at 70 degrees C increased around four times, while half-life in 80% (v/v) dioxane increased 22 times. After repeated use of 6 times in batch reactor for pyrogallol oxidation immobilized HRP retained 45% of original activity.
PB  - Elsevier
T2  - Reactive and Functional Polymers
T1  - Tyramine modified alginates via periodate oxidation for peroxidase induced hydrogel formation and immobilization
VL  - 93
SP  - 77
EP  - 83
DO  - 10.1016/j.reactfunctpolym.2015.06.004
ER  - 

@article{
author = "Prodanović, Olivera and Spasojevic, Dragica and Prokopijevic, Milos and Radotić, Ksenija and Marković, Nevena and Blažić, Marija and Prodanović, Radivoje",
year = "2015",
abstract = "Phenol and amino groups were introduced into alginate to different degrees via oxidation with 2.5, 5, 10, 15 and 20 mol% of periodate and reductive amination by tyramine. Modification of alginate with tyramine was confirmed by FTIR spectroscopy and UV-VIS spectroscopy, while concentration of phenol and ionizable groups was determined using absorbance at 275 nm and acid-base titration. All tyramine-alginates were able to form hydrogels after cross-linking with horse radish peroxidase (HRP) and hydrogen peroxide. Tyramine-alginates oxidized with up to 10 mol% of periodate were also capable of forming hydrogels with calcium ions. Tyramine-alginates were tested for HRP immobilization within micro-beads obtained by peroxidase catalyzed droplet polymerization using internal delivery of hydrogen peroxide via glucose oxidase and glucose. Highest activity of immobilized peroxidase was obtained with 20% (w/v) tyramine-alginate obtained via 20 mol% periodate oxidation. Immobilized enzyme was not leaking from the micro-beads and was further kinetically characterized for pyrogallol oxidation. Km for pyrogallol was increased after immobilization from 1.93 mM for soluble HRP to 734 mM for immobilized HRP. The optimum pH was also increased from pH 7.0 to 8.0. Temperature and organic solvent stability improved significantly after immobilization, so that half-life at 70 degrees C increased around four times, while half-life in 80% (v/v) dioxane increased 22 times. After repeated use of 6 times in batch reactor for pyrogallol oxidation immobilized HRP retained 45% of original activity.",
publisher = "Elsevier",
journal = "Reactive and Functional Polymers",
title = "Tyramine modified alginates via periodate oxidation for peroxidase induced hydrogel formation and immobilization",
volume = "93",
pages = "77-83",
doi = "10.1016/j.reactfunctpolym.2015.06.004"
}

Prodanović, O., Spasojevic, D., Prokopijevic, M., Radotić, K., Marković, N., Blažić, M.,& Prodanović, R.. (2015). Tyramine modified alginates via periodate oxidation for peroxidase induced hydrogel formation and immobilization. in Reactive and Functional Polymers
Elsevier., 93, 77-83.
https://doi.org/10.1016/j.reactfunctpolym.2015.06.004

Prodanović O, Spasojevic D, Prokopijevic M, Radotić K, Marković N, Blažić M, Prodanović R. Tyramine modified alginates via periodate oxidation for peroxidase induced hydrogel formation and immobilization. in Reactive and Functional Polymers. 2015;93:77-83.
doi:10.1016/j.reactfunctpolym.2015.06.004 .

Prodanović, Olivera, Spasojevic, Dragica, Prokopijevic, Milos, Radotić, Ksenija, Marković, Nevena, Blažić, Marija, Prodanović, Radivoje, "Tyramine modified alginates via periodate oxidation for peroxidase induced hydrogel formation and immobilization" in Reactive and Functional Polymers, 93 (2015):77-83,
https://doi.org/10.1016/j.reactfunctpolym.2015.06.004 . .

TY  - JOUR
AU  - Grozdanović, Milica
AU  - Ostojić, Sanja
AU  - Aleksić, Ivana
AU  - Anđelković, Uroš
AU  - Petersen, Arnd
AU  - Gavrović-Jankulović, Marija
PY  - 2014
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/1397
AB  - BACKGROUND: Actinidin is a cysteine protease and major allergen from kiwi fruit. When purified under specific native conditions, actinidin preparations from fresh kiwi fruit contain both an active and inactive form of this enzyme. In this study, biochemical and immunological properties upon simulated gastro-intestinal digestion, as well as thermal stability, were investigated for both active and E-64-inhibited actinidin. RESULTS: Active actinidin retained its primary structure and proteolytic activity after 2 h of simulated gastric digestion, followed by 2 h of intestinal digestion, as assessed by SDS-PAGE, zymography and mass spectroscopy. Immunological reactivity of active actinidin was also preserved, as tested by immunoelectrophoresis. The E-64 inhibited actinidin was fully degraded after 1 h of pepsin treatment. Differential scanning calorimetry showed that active actinidin has one transition maximum temperature (T-m) at 73.9 degrees C, whereas in the E-64-actinidin complex the two actinidin domains unfolded independently, with the first domain having a T-m value of only 61 degrees C. CONCLUSION: Active actinidin is capable of reaching the intestinal mucosa in a proteolytically active and immunogenic state. Inhibitor binding induces changes in the actinidin molecule that go beyond inhibition of proteolytic activity, also influencing the digestion stability and T-m values of actinidin, features important in the characterisation of food allergens.
PB  - Wiley-Blackwell, Hoboken
T2  - Journal of the Science of Food and Agriculture
T1  - Active actinidin retains function upon gastro-intestinal digestion and is more thermostable than the E-64-inhibited counterpart
VL  - 94
IS  - 14
SP  - 3046
EP  - 3052
DO  - 10.1002/jsfa.6656
ER  - 

@article{
author = "Grozdanović, Milica and Ostojić, Sanja and Aleksić, Ivana and Anđelković, Uroš and Petersen, Arnd and Gavrović-Jankulović, Marija",
year = "2014",
abstract = "BACKGROUND: Actinidin is a cysteine protease and major allergen from kiwi fruit. When purified under specific native conditions, actinidin preparations from fresh kiwi fruit contain both an active and inactive form of this enzyme. In this study, biochemical and immunological properties upon simulated gastro-intestinal digestion, as well as thermal stability, were investigated for both active and E-64-inhibited actinidin. RESULTS: Active actinidin retained its primary structure and proteolytic activity after 2 h of simulated gastric digestion, followed by 2 h of intestinal digestion, as assessed by SDS-PAGE, zymography and mass spectroscopy. Immunological reactivity of active actinidin was also preserved, as tested by immunoelectrophoresis. The E-64 inhibited actinidin was fully degraded after 1 h of pepsin treatment. Differential scanning calorimetry showed that active actinidin has one transition maximum temperature (T-m) at 73.9 degrees C, whereas in the E-64-actinidin complex the two actinidin domains unfolded independently, with the first domain having a T-m value of only 61 degrees C. CONCLUSION: Active actinidin is capable of reaching the intestinal mucosa in a proteolytically active and immunogenic state. Inhibitor binding induces changes in the actinidin molecule that go beyond inhibition of proteolytic activity, also influencing the digestion stability and T-m values of actinidin, features important in the characterisation of food allergens.",
publisher = "Wiley-Blackwell, Hoboken",
journal = "Journal of the Science of Food and Agriculture",
title = "Active actinidin retains function upon gastro-intestinal digestion and is more thermostable than the E-64-inhibited counterpart",
volume = "94",
number = "14",
pages = "3046-3052",
doi = "10.1002/jsfa.6656"
}

Grozdanović, M., Ostojić, S., Aleksić, I., Anđelković, U., Petersen, A.,& Gavrović-Jankulović, M.. (2014). Active actinidin retains function upon gastro-intestinal digestion and is more thermostable than the E-64-inhibited counterpart. in Journal of the Science of Food and Agriculture
Wiley-Blackwell, Hoboken., 94(14), 3046-3052.
https://doi.org/10.1002/jsfa.6656

Grozdanović M, Ostojić S, Aleksić I, Anđelković U, Petersen A, Gavrović-Jankulović M. Active actinidin retains function upon gastro-intestinal digestion and is more thermostable than the E-64-inhibited counterpart. in Journal of the Science of Food and Agriculture. 2014;94(14):3046-3052.
doi:10.1002/jsfa.6656 .

Grozdanović, Milica, Ostojić, Sanja, Aleksić, Ivana, Anđelković, Uroš, Petersen, Arnd, Gavrović-Jankulović, Marija, "Active actinidin retains function upon gastro-intestinal digestion and is more thermostable than the E-64-inhibited counterpart" in Journal of the Science of Food and Agriculture, 94, no. 14 (2014):3046-3052,
https://doi.org/10.1002/jsfa.6656 . .

TY  - JOUR
AU  - Kovačević, Gordana
AU  - Blažić, Marija
AU  - Draganic, Bojana
AU  - Ostafe, Raluca
AU  - Gavrović-Jankulović, Marija
AU  - Fischer, Rainer
AU  - Prodanović, Radivoje
PY  - 2014
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/1506
AB  - Aspergillus niger glucose oxidase (GOx) genes for wild-type (GenBank accession no. X16061, swiss-Prot; P13006) and M12 mutant (N2Y, K13E, T30 V, I94 V, K152R) were cloned into pPICZ alpha A vector for expression in Pichia pastoris KM71H strain. The highest expression level of 17.5 U/mL of fermentation media was obtained in 0.5 % (v/v) methanol after 9 days of fermentation. The recombinant GOx was purified by cross-flow ultrafiltration using membranes of 30 kDa molecular cutoff and DEAE ion-exchange chromatography at pH 6.0. Purified wt GOx had k (cat) of 189.4 s(-1) and K (m) of 28.26 mM while M12 GOx had k (cat) of 352.0 s(-1) and K (m) of 13.33 mM for glucose at pH 5.5. Specificity constants k (cat)/K (m) of wt (6.70 mM(-1) s(-1)) and M12 GOx (26.7 mM(-1) s(-1)) expressed in P. pastoris KM71H were around three times higher than for the same enzymes previously expressed in Saccharomyces cerevisiae InvSc1 strain. The pH optimum and sugar specificity of M12 mutant of GOx remained similar to the wild-type form of the enzyme, while thermostability was slightly decreased. M12 GOx expressed in P. pastoris showed three times higher activity compared to the wt GOx toward redox mediators like N,N-dimethyl-nitroso-aniline used for glucose strips manufacturing. M12 mutant of GOx produced in P. pastoris KM71H could be useful for manufacturing of glucose biosensors and biofuel cells.
PB  - Humana Press Inc, Totowa
T2  - Molecular Biotechnology
T1  - Cloning, Heterologous Expression, Purification and Characterization of M12 Mutant of Aspergillus niger Glucose Oxidase in Yeast Pichia pastoris KM71H
VL  - 56
IS  - 4
SP  - 305
EP  - 311
DO  - 10.1007/s12033-013-9709-x
ER  - 

@article{
author = "Kovačević, Gordana and Blažić, Marija and Draganic, Bojana and Ostafe, Raluca and Gavrović-Jankulović, Marija and Fischer, Rainer and Prodanović, Radivoje",
year = "2014",
abstract = "Aspergillus niger glucose oxidase (GOx) genes for wild-type (GenBank accession no. X16061, swiss-Prot; P13006) and M12 mutant (N2Y, K13E, T30 V, I94 V, K152R) were cloned into pPICZ alpha A vector for expression in Pichia pastoris KM71H strain. The highest expression level of 17.5 U/mL of fermentation media was obtained in 0.5 % (v/v) methanol after 9 days of fermentation. The recombinant GOx was purified by cross-flow ultrafiltration using membranes of 30 kDa molecular cutoff and DEAE ion-exchange chromatography at pH 6.0. Purified wt GOx had k (cat) of 189.4 s(-1) and K (m) of 28.26 mM while M12 GOx had k (cat) of 352.0 s(-1) and K (m) of 13.33 mM for glucose at pH 5.5. Specificity constants k (cat)/K (m) of wt (6.70 mM(-1) s(-1)) and M12 GOx (26.7 mM(-1) s(-1)) expressed in P. pastoris KM71H were around three times higher than for the same enzymes previously expressed in Saccharomyces cerevisiae InvSc1 strain. The pH optimum and sugar specificity of M12 mutant of GOx remained similar to the wild-type form of the enzyme, while thermostability was slightly decreased. M12 GOx expressed in P. pastoris showed three times higher activity compared to the wt GOx toward redox mediators like N,N-dimethyl-nitroso-aniline used for glucose strips manufacturing. M12 mutant of GOx produced in P. pastoris KM71H could be useful for manufacturing of glucose biosensors and biofuel cells.",
publisher = "Humana Press Inc, Totowa",
journal = "Molecular Biotechnology",
title = "Cloning, Heterologous Expression, Purification and Characterization of M12 Mutant of Aspergillus niger Glucose Oxidase in Yeast Pichia pastoris KM71H",
volume = "56",
number = "4",
pages = "305-311",
doi = "10.1007/s12033-013-9709-x"
}

Kovačević, G., Blažić, M., Draganic, B., Ostafe, R., Gavrović-Jankulović, M., Fischer, R.,& Prodanović, R.. (2014). Cloning, Heterologous Expression, Purification and Characterization of M12 Mutant of Aspergillus niger Glucose Oxidase in Yeast Pichia pastoris KM71H. in Molecular Biotechnology
Humana Press Inc, Totowa., 56(4), 305-311.
https://doi.org/10.1007/s12033-013-9709-x

Kovačević G, Blažić M, Draganic B, Ostafe R, Gavrović-Jankulović M, Fischer R, Prodanović R. Cloning, Heterologous Expression, Purification and Characterization of M12 Mutant of Aspergillus niger Glucose Oxidase in Yeast Pichia pastoris KM71H. in Molecular Biotechnology. 2014;56(4):305-311.
doi:10.1007/s12033-013-9709-x .

Kovačević, Gordana, Blažić, Marija, Draganic, Bojana, Ostafe, Raluca, Gavrović-Jankulović, Marija, Fischer, Rainer, Prodanović, Radivoje, "Cloning, Heterologous Expression, Purification and Characterization of M12 Mutant of Aspergillus niger Glucose Oxidase in Yeast Pichia pastoris KM71H" in Molecular Biotechnology, 56, no. 4 (2014):305-311,
https://doi.org/10.1007/s12033-013-9709-x . .

TY  - JOUR
AU  - Prokopovic, Vladimir
AU  - Popović, Milica
AU  - Anđelković, Uroš
AU  - Marsavelski, Aleksandra
AU  - Raskovic, Brankica
AU  - Gavrović-Jankulović, Marija
AU  - Polović, Natalija
PY  - 2014
UR  - https://cer.ihtm.bg.ac.rs/handle/123456789/1508
AB  - Objective: Human BPIFA2 (parotid secretory protein) is a ubiquitous soluble salivary protein, which belongs to the PLUNC family of proteins. Having sequence similarity to bactericidal/permeability-increasing protein and lipopolysaccharide-binding protein, PLUNC proteins are probably involved in local antibacterial response at mucosal sites, such as oral cavity. The aim of the study was to isolate and characterize human BPIFA2. Design: In this paper, we report one-step affinity chromatography method for BPIFA2 purification from whole human saliva. The isolated BPIFA2 was identified by trypsin mass fingerprinting and characterized by electrophoretic methods. Antibacterial activity of BPIFA2 against model microorganism Pseudomonas aeruginosa was shown in minimum inhibitory concentration and time kill study assays. Results: The protein showed microheterogeneity, both in molecular weight and pI value. BPIFA2 inhibited the growth of P. aeruginosa in microgram concentration range determined by minimum inhibitory concentration assay. In the time kill study, 32 mu g/mL BPIFA2 showed clear bactericidal activity and did not cause any aggregation of bacteria. Conclusion: Affinity chromatography is well suited for isolation of functional BPIFA2 with a potent bactericidal activity against P. aeruginosa.
PB  - Oxford : Pergamon-Elsevier Science Ltd
T2  - Archives of Oral Biology
T1  - Isolation, biochemical characterization and anti-bacterial activity of BPIFA2 protein
VL  - 59
IS  - 3
SP  - 302
EP  - 309
DO  - 10.1016/j.archoralbio.2013.12.005
ER  - 

@article{
author = "Prokopovic, Vladimir and Popović, Milica and Anđelković, Uroš and Marsavelski, Aleksandra and Raskovic, Brankica and Gavrović-Jankulović, Marija and Polović, Natalija",
year = "2014",
abstract = "Objective: Human BPIFA2 (parotid secretory protein) is a ubiquitous soluble salivary protein, which belongs to the PLUNC family of proteins. Having sequence similarity to bactericidal/permeability-increasing protein and lipopolysaccharide-binding protein, PLUNC proteins are probably involved in local antibacterial response at mucosal sites, such as oral cavity. The aim of the study was to isolate and characterize human BPIFA2. Design: In this paper, we report one-step affinity chromatography method for BPIFA2 purification from whole human saliva. The isolated BPIFA2 was identified by trypsin mass fingerprinting and characterized by electrophoretic methods. Antibacterial activity of BPIFA2 against model microorganism Pseudomonas aeruginosa was shown in minimum inhibitory concentration and time kill study assays. Results: The protein showed microheterogeneity, both in molecular weight and pI value. BPIFA2 inhibited the growth of P. aeruginosa in microgram concentration range determined by minimum inhibitory concentration assay. In the time kill study, 32 mu g/mL BPIFA2 showed clear bactericidal activity and did not cause any aggregation of bacteria. Conclusion: Affinity chromatography is well suited for isolation of functional BPIFA2 with a potent bactericidal activity against P. aeruginosa.",
publisher = "Oxford : Pergamon-Elsevier Science Ltd",
journal = "Archives of Oral Biology",
title = "Isolation, biochemical characterization and anti-bacterial activity of BPIFA2 protein",
volume = "59",
number = "3",
pages = "302-309",
doi = "10.1016/j.archoralbio.2013.12.005"
}

Prokopovic, V., Popović, M., Anđelković, U., Marsavelski, A., Raskovic, B., Gavrović-Jankulović, M.,& Polović, N.. (2014). Isolation, biochemical characterization and anti-bacterial activity of BPIFA2 protein. in Archives of Oral Biology
Oxford : Pergamon-Elsevier Science Ltd., 59(3), 302-309.
https://doi.org/10.1016/j.archoralbio.2013.12.005

Prokopovic V, Popović M, Anđelković U, Marsavelski A, Raskovic B, Gavrović-Jankulović M, Polović N. Isolation, biochemical characterization and anti-bacterial activity of BPIFA2 protein. in Archives of Oral Biology. 2014;59(3):302-309.
doi:10.1016/j.archoralbio.2013.12.005 .

Prokopovic, Vladimir, Popović, Milica, Anđelković, Uroš, Marsavelski, Aleksandra, Raskovic, Brankica, Gavrović-Jankulović, Marija, Polović, Natalija, "Isolation, biochemical characterization and anti-bacterial activity of BPIFA2 protein" in Archives of Oral Biology, 59, no. 3 (2014):302-309,
https://doi.org/10.1016/j.archoralbio.2013.12.005 . .