The role of Tyr102 residue in the functioning of bacterial NAD+-dependent formate dehydrogenase of Pseudomonas sp. 101

Popinako, A.V.; Pometun, A.A.; Nilov, D.K.; Dibrova, D.V.; Khrustalev, V.V.; Khrustaleva, T.A.; Iurchenko, T.S.; Nikolaeva, A.Y.; Švedas, V.K.; Boyko, K.M.; Tishkov, V.I.; Popov, V.O.

Авторы: Popinako A.V., Pometun A.A., Nilov D.K., Dibrova D.V., Khrustalev V.V., Khrustaleva T.A., Iurchenko T.S., Nikolaeva A.Yu, Švedas V.K., Boyko K.M., Tishkov V.I., Popov V.O.
Журнал: Biochemical and Biophysical Research Communications
Том: 616
Год издания: 2022
Издательство: Academic Press
Местоположение издательства: United States
Первая страница: 134
Последняя страница: 139
DOI: 10.1016/j.bbrc.2022.05.064
Аннотация: Formate oxidation is a final step of methanol oxidation in methylotrophic prokaryotes and is important for detoxification of formate in other organisms. The structural mechanism of the formate dehydrogenase (FDH) of Pseudomonas sp. 101 has been studied for about 30 years. In the active center of FDH, the oxidation of formic acid into carbon dioxide in a NADþ-dependent way takes place. Residues that form the active center of that enzyme, as well as those that form the so-called substrate channel, are engagedin the catalytic cycle. Our study allowed to characterize a new residue, Tyr102, involved in the work ofthe enzyme. This residue is located in the outer neck of the substrate channel (at the beginning of the path of the substrate to the active center) and acts as a “button” which connects two enzyme domainsinto an active, “buttoned up” conformation. Our study of the kinetic parameters of mutant enzymes hasshown that Tyr102Phe substitution leads to an approximately 80-fold increase of the Michaelis constantrelative to the native enzyme, unlike Phe311Trp and Phe311Tyr substitution of neighboring residuePhe311. Our analysis of the Tyr102Phe mutant in the open conformation by X-ray crystallography hasshown that its overall fold remains almost the same as that of the native enzyme. Molecular dynamicssimulations of the ternary complexes of the native FDH enzyme and its Tyr102Phe mutant showed thatTyr102Phe substitution results in the loss of an interdomain hydrogen bond between the Tyr102 andGln313 residues, which, in turn, destabilizes the closed conformation and affects the isolation of the FDH active site from water molecules. Our structural investigations have shown that Tyr102Phe replacement also leads to the destruction of interdomain contacts of Phe102 with Phe311, Pro312 residues, and decreases the stability of the Leu103-Val127 beta bridge. Phylogenetic analysis also confirmed the importance of the Tyr102 residue for enzymes from the FDH family, in which it is absolutely conserved.
Добавил в систему: Тишков Владимир Иванович

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The role of Tyr102 residue in the functioning of bacterial NAD+-dependent formate dehydrogenase of Pseudomonas sp. 101статья

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