title: Asp477 is a determinant of the enantioselectivity in yeast transketolase.
abstract: The conserved residue Asp477 in yeast transketolase is located in the substrate channel of the enzyme and forms a hydrogen bond with the C2-hydroxyl group of the acceptor substrate. The significance of this interaction for the recognition of the preferred acceptor substrates, D-alpha-hydroxyaldehydes was investigated by site-directed mutagenesis. In the wild-type enzyme the kcat/KM values are by three to four orders of magnitude lower for 2-deoxyaldoses or substrates with L-configuration at the C2-atom. In the Asp477 Ala mutant, the kcat/KM values for D-alpha-hydroxyaldehydes are decreased by a thousandfold, while the kcat/KM values for substrates with L-configuration or 2-deoxyaldoses are similar to wild-type enzyme. These results indicate that Asp477 is involved in determining the enantioselectivity of transketolase.
authors.affiliation: Department of Medical Biochemistry and Biophysics, Karolinska Institutet, Stockholm, Sweden.
authors.forename: U
authors.initials: U
authors.lastname: Nilsson
authors.forename: L
authors.initials: L
authors.lastname: Hecquet
authors.forename: T
authors.initials: T
authors.lastname: Gefflaut
authors.forename: C
authors.initials: C
authors.lastname: Guerard
authors.forename: G
authors.initials: G
authors.lastname: Schneider
mesh_terms: D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D013379:Substrate Specificity; D014174:Transketolase / Q000235:genetics / Q000378:metabolism*; D015003:Yeasts / Q000201:enzymology*
chemical_list: D014174:Transketolase