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In search for structural targets for engineering D-amino acid transaminase: modulation of pH optimum and substrate specificityстатья
Информация о цитировании статьи получена из
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Статья опубликована в журнале из списка Web of Science и/или Scopus
Дата последнего поиска статьи во внешних источниках: 20 февраля 2024 г.
- Авторы: Shilova S.A., Matyuta I.O., Khrenova M.G., Nikolaeva A.Y., Klyachko N.L., Minyaev M.E., Khomutov A.R., Boyko K.M., Popov V.O., Bezsudnova E.Y.
- Журнал: Biochemical Journal
- Том: 480
- Номер: 16
- Год издания: 2023
- Издательство: Portland Press, Ltd.
- Местоположение издательства: United Kingdom
- Первая страница: 1267
- Последняя страница: 1284
- DOI: 10.1042/BCJ20230233
- Аннотация: Shilova, S.A.; Matyuta, I.O.; Khrenova, M.G.; Nikolaeva, A.Y.; Klyachko, N.L.; Minyaev, M.E.; Khomutov, A.R.; Boyko, K.M.; Popov, V.O.; Bezsudnova, E.Y. (2023). In search for structural targets for engineering D-amino acid transaminase: modulation of pH optimum and substrate specificity. Biochemical Journal. V. 480. P.1267–1284. https://doi.org/10.1042/BCJ20230233 (IF 3.87 SJR Q1 Biochemistry), Published 23 August 2023The development of biocatalysts requires reorganization of the enzyme’s active site tofacilitate the productive binding of the target substrate and improve turnover number atdesired conditions. Pyridoxal-50-phosphate (PLP) - dependent transaminases are highlyefficient biocatalysts for asymmetric amination of ketones and keto acids. However,transaminases, being stereoselective enzymes, have a narrow substrate specificity due tothe ordered structure of the active site and work only in neutral-alkaline media. Here, weinvestigated the D-amino acid transaminase from Aminobacterium colombiense, with theactive site organized differently from that of the canonical D-amino acid transaminasefrom Bacillus sp. YM-1. Using a combination of site-directed mutagenesis, kinetic analysis,molecular modeling, and structural analysis we determined the active site residuesresponsible for substrate binding, substrate differentiation, thermostability of a functionaldimer, and affecting the pH optimum. We demonstrated that the high specificity towardD-glutamate/α-ketoglutarate is due to the interactions of a γ-carboxylate group with K237residue, while binding of other substrates stems from the effectiveness of their accommodationin the active site optimized for D-glutamate/α-ketoglutarate binding. Furthermore,we showed that the K237A substitution shifts the catalytic activity optimum to acidic pH.Our findings are useful for achieving target substrate specificity and demonstrate thepotential for developing and optimizing transaminases for various applications.Introduction
- Добавил в систему: Клячко Наталья Львовна