Heterologous Expression and Partial Characterization of a Putative Opine Dehydrogenase from a Metagenomic Sequence of Desulfohalobium retbaense
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2023
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The aim of this research was to prove the function of the putative opine dehydrogenase from Desulfohalobium retbaense and to characterize the enzyme in terms of functional and kinetic parameters. A putative opine dehydrogenase was identified from a metagenomic library by a sequence-based technique search of the metagenomic library, and afterward was successfully heterologously produced in Escherichia coli. In order to examine its potential for applications in the synthesis of secondary amines, first the substrate specificity of the enzyme towards different amino donors and amino acceptors was determined. The highest affinity was observed towards small amino acids, preferentially L-alanine, and when it comes to α-keto acids, pyruvate proved to be a preferential amino acceptor. The highest activity was observed at pH 6.5 in the absence of salts. The enzyme showed remarkable stability in a wide range of experimental conditions, such as broad pH stability (from 6.0–11.0 after 30 min incuba...tion in buffers at a certain pH), stability in the presence of NaCl up to 3.0 M for 24 h, it retained 80% of the initial activity after 1 h incubation at 45°C, and 65% of the initial activity after 24 h incubation in 30% dimethyl sulfoxide.
Ključne reči:
biocatalysis / chiral amines / halophiles / metagenomics / protein expressionIzvor:
ChemBioChem, 2023, 24, 20Izdavač:
- Wiley-VCH GmbH
Finansiranje / projekti:
- Ministarstvo nauke, tehnološkog razvoja i inovacija Republike Srbije, institucionalno finansiranje - 200026 (Univerzitet u Beogradu, Institut za hemiju, tehnologiju i metalurgiju - IHTM) (RS-MESTD-inst-2020-200026)
- Ministarstvo nauke, tehnološkog razvoja i inovacija Republike Srbije, institucionalno finansiranje - 200168 (Univerzitet u Beogradu, Hemijski fakultet) (RS-MESTD-inst-2020-200168)
- Ministarstvo nauke, tehnološkog razvoja i inovacija Republike Srbije, institucionalno finansiranje - 200053 (Univerzitet u Beogradu, Institut za multidisciplinarna istraživanja) (RS-MESTD-inst-2020-200053)
DOI: 10.1002/cbic.202300414
ISSN: 1439-4227
WoS: 001052168000001
Scopus: 2-s2.0-85168572903
Institucija/grupa
IHTMTY - JOUR AU - Kaličanin, Nevena AU - Balaž, Ana Marija AU - Prodanović, Olivera AU - Prodanović, Radivoje PY - 2023 UR - https://cer.ihtm.bg.ac.rs/handle/123456789/7171 AB - The aim of this research was to prove the function of the putative opine dehydrogenase from Desulfohalobium retbaense and to characterize the enzyme in terms of functional and kinetic parameters. A putative opine dehydrogenase was identified from a metagenomic library by a sequence-based technique search of the metagenomic library, and afterward was successfully heterologously produced in Escherichia coli. In order to examine its potential for applications in the synthesis of secondary amines, first the substrate specificity of the enzyme towards different amino donors and amino acceptors was determined. The highest affinity was observed towards small amino acids, preferentially L-alanine, and when it comes to α-keto acids, pyruvate proved to be a preferential amino acceptor. The highest activity was observed at pH 6.5 in the absence of salts. The enzyme showed remarkable stability in a wide range of experimental conditions, such as broad pH stability (from 6.0–11.0 after 30 min incubation in buffers at a certain pH), stability in the presence of NaCl up to 3.0 M for 24 h, it retained 80% of the initial activity after 1 h incubation at 45°C, and 65% of the initial activity after 24 h incubation in 30% dimethyl sulfoxide. PB - Wiley-VCH GmbH T2 - ChemBioChem T1 - Heterologous Expression and Partial Characterization of a Putative Opine Dehydrogenase from a Metagenomic Sequence of Desulfohalobium retbaense VL - 24 IS - 20 IS - e202300414 DO - 10.1002/cbic.202300414 ER -
@article{ author = "Kaličanin, Nevena and Balaž, Ana Marija and Prodanović, Olivera and Prodanović, Radivoje", year = "2023", abstract = "The aim of this research was to prove the function of the putative opine dehydrogenase from Desulfohalobium retbaense and to characterize the enzyme in terms of functional and kinetic parameters. A putative opine dehydrogenase was identified from a metagenomic library by a sequence-based technique search of the metagenomic library, and afterward was successfully heterologously produced in Escherichia coli. In order to examine its potential for applications in the synthesis of secondary amines, first the substrate specificity of the enzyme towards different amino donors and amino acceptors was determined. The highest affinity was observed towards small amino acids, preferentially L-alanine, and when it comes to α-keto acids, pyruvate proved to be a preferential amino acceptor. The highest activity was observed at pH 6.5 in the absence of salts. The enzyme showed remarkable stability in a wide range of experimental conditions, such as broad pH stability (from 6.0–11.0 after 30 min incubation in buffers at a certain pH), stability in the presence of NaCl up to 3.0 M for 24 h, it retained 80% of the initial activity after 1 h incubation at 45°C, and 65% of the initial activity after 24 h incubation in 30% dimethyl sulfoxide.", publisher = "Wiley-VCH GmbH", journal = "ChemBioChem", title = "Heterologous Expression and Partial Characterization of a Putative Opine Dehydrogenase from a Metagenomic Sequence of Desulfohalobium retbaense", volume = "24", number = "20, e202300414", doi = "10.1002/cbic.202300414" }
Kaličanin, N., Balaž, A. M., Prodanović, O.,& Prodanović, R.. (2023). Heterologous Expression and Partial Characterization of a Putative Opine Dehydrogenase from a Metagenomic Sequence of Desulfohalobium retbaense. in ChemBioChem Wiley-VCH GmbH., 24(20). https://doi.org/10.1002/cbic.202300414
Kaličanin N, Balaž AM, Prodanović O, Prodanović R. Heterologous Expression and Partial Characterization of a Putative Opine Dehydrogenase from a Metagenomic Sequence of Desulfohalobium retbaense. in ChemBioChem. 2023;24(20). doi:10.1002/cbic.202300414 .
Kaličanin, Nevena, Balaž, Ana Marija, Prodanović, Olivera, Prodanović, Radivoje, "Heterologous Expression and Partial Characterization of a Putative Opine Dehydrogenase from a Metagenomic Sequence of Desulfohalobium retbaense" in ChemBioChem, 24, no. 20 (2023), https://doi.org/10.1002/cbic.202300414 . .