Asp477 is a determinant of the enantioselectivity in yeast transketolase.
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.
Nilsson
U
U
Department of Medical Biochemistry and Biophysics, Karolinska Institutet, Stockholm, Sweden.
Hecquet
L
L
Gefflaut
T
T
Guerard
C
C
Schneider
G
G
D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D013379:Substrate Specificity; D014174:Transketolase / Q000235:genetics / Q000378:metabolism*; D015003:Yeasts / Q000201:enzymology*
D014174:Transketolase