Role of the histidine 176 residue in glyceraldehyde-3-phosphate dehydrogenase as probed by site-directed mutagenesis.
The catalytically essential amino acid, histidine 176, in the active site of Escherichia coli glyceraldehyde-3-phosphate dehydrogenase (GAPDH) has been replaced with an asparagine residue by site-directed mutagenesis. The role of histidine 176 as a chemical activator, enhancing the reactivity of the thiol group of cysteine 149, has been demonstrated, with iodoacetamide as a probe. The esterolytic properties of GAPDH, illustrated by the hydrolysis of p-nitrophenyl acetate, have been also studied. The kinetic results favor a role for histidine 176 not only as a chemical activator of cysteine 149 but also as a hydrogen donor facilitating the formation of tetrahedral intermediates. These results support the hypothesis that histidine 176 plays a similar role during the oxidative phosphorylation of glyceraldehyde 3-phosphate.
Soukri
A
A
Laboratoire d'Enzymologie et de Génie Génétique, UA CNRS, 457, Faculté des Sciences, Vandoeuvre les Nancy, France.
Mougin
A
A
Corbier
C
C
Wonacott
A
A
Branlant
C
C
Branlant
G
G
D001665:Binding Sites; D004926:Escherichia coli / Q000201:enzymology*; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000235:genetics / Q000378:metabolism*; D006639:Histidine; D007700:Kinetics; D008433:Mathematics; D008962:Models, Theoretical; D009154:Mutation
D006639:Histidine; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases