title: Stereochemical course of hydrolytic reaction catalyzed by alpha-galactosidase from cold adaptable marine bacterium of genus Pseudoalteromonas.
abstract: The recombinant α-galactosidase of the marine bacterium (α-PsGal) was synthesized with the use of the plasmid 40Gal, consisting of plasmid pET-40b (+) (Novagen) and the gene corresponding to the open reading frame of the mature α-galactosidase of marine bacterium Pseudoalteromonas sp. KMM 701, transformed into the Escherichia coli Rosetta(DE3) cells. In order to understand the mechanism of action, the stereochemistry of hydrolysis of 4-nitrophenyl α-D-galactopyranoside (4-NPGP) by α-PsGal was measured by (1)H NMR spectroscopy. The kinetics of formation of α- and β-anomer of galactose showed that α-anomer initially formed and accumulated, and then an appreciable amount of β-anomer appeared as a result of mutarotation. The data clearly show that the enzymatic hydrolysis of 4-NPGP proceeds with the retention of anomeric configuration, probably, due to a double displacement mechanism of reaction.
authors.affiliation: G.B. Elyakov Pacific Institute of Bioorganic Chemistry, Far Eastern Branch, Russian Academy of Sciences Vladivostok, Russia.
authors.forename: Irina Y
authors.initials: IY
authors.lastname: Bakunina
authors.affiliation: G.B. Elyakov Pacific Institute of Bioorganic Chemistry, Far Eastern Branch, Russian Academy of Sciences Vladivostok, Russia.
authors.forename: Larissa A
authors.initials: LA
authors.lastname: Balabanova
authors.affiliation: G.B. Elyakov Pacific Institute of Bioorganic Chemistry, Far Eastern Branch, Russian Academy of Sciences Vladivostok, Russia.
authors.forename: Vasiliy A
authors.initials: VA
authors.lastname: Golotin
authors.affiliation: G.B. Elyakov Pacific Institute of Bioorganic Chemistry, Far Eastern Branch, Russian Academy of Sciences Vladivostok, Russia.
authors.forename: Lyubov V
authors.initials: LV
authors.lastname: Slepchenko
authors.affiliation: G.B. Elyakov Pacific Institute of Bioorganic Chemistry, Far Eastern Branch, Russian Academy of Sciences Vladivostok, Russia.
authors.forename: Vladimir V
authors.initials: VV
authors.lastname: Isakov
authors.affiliation: G.B. Elyakov Pacific Institute of Bioorganic Chemistry, Far Eastern Branch, Russian Academy of Sciences Vladivostok, Russia.
authors.forename: Valeriy A
authors.initials: VA
authors.lastname: Rasskazov
keywords: 1H NMR spectroscopy; Recombinant retaining α-galactosidase; marine bacterium