Effect of the Components of a Buffer Solution on the Catalytic Activity of the NAD+-Dependent Formate Dehydrogenase from the Bacterium Staphylococcus aureus

Iurchenko, T.S.; Bolotova, S.B.; Loginova, A.A.; Pometun, E.V.; Savin, S.S.; Pometun, A.A.; Tishkov, V.I.

Авторы: Iurchenko T.S., Bolotova S.B., Loginova A.A., Pometun E.V., Savin S.S., Pometun A.A., Tishkov V.I.
Журнал: Moscow University Chemistry Bulletin
Том: 77
Номер: 6
Год издания: 2022
Издательство: Allerton Press Inc.
Местоположение издательства: United States
Первая страница: 330
Последняя страница: 339
DOI: 10.3103/s0027131422060074
Аннотация: NAD+-dependent formate dehydrogenase (FDH, EC 1.2.1.2) from bacterium Staphylococcus aureus (SauFDH) is the most active enzyme among FDHs of this group, but high values of KM of the enzyme with NAD+ and formate in standard 0.1 M phosphate buffer result in lower catalytic efficiency kcat/KM compared to ones for other FDHs. We have studied influence of different buffers on catalytic properties of SauFDH. Sodium phosphate (NaPB) was used as the base buffer component and Tris, Gly and citrate (Cit) were added to NaPB to prepare double, ternary and quaternary buffer systems with different concentrations. It was found that KM for formate does not depend on buffer composition and concentration, while values of kcat и KMNAD+ increased and decreased significantly. The highest positive effect achieved in the case of quaternary buffer NaPB-Cit-Tris-Gly. At 0.05 M concentration of each component kcat increased by 70% compared to one in standard 0.1 M NaPB. At 0.1 M of each component improvement of both parameters, kcat and KMNAD+ was observed. Thermal inactivation studies in NaPB and complex NaPB-Cit-Tris-Gly buffer showed that at component concentrations 0.1 M and more SauFDH thermal stability increased. Value of stabilization effect depends on ion strength but not on type of buffer. Comparison of X-ray structures of holo-forms of SauFDH and FDH from bacterium Pseudomonas sp.101 shows that active site of PseFDH in complex with substrate is totally closed, while in holo-SauFDH amino acid residues in active site can be accessed by water molecules and buffer components. It could be the reason of kcat и KMNAD+ changes in buffers of different compositions.
Добавил в систему: Тишков Владимир Иванович

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Effect of the Components of a Buffer Solution on the Catalytic Activity of the NAD+-Dependent Formate Dehydrogenase from the Bacterium Staphylococcus aureusстатья

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