The 2'-phosphate of NADP is critical for optimum productive binding to 6-phosphogluconate dehydrogenase from Candida utilis.
Initial velocity studies obtained with alternative dinucleotide substrates for the 6-phosphogluconate dehydrogenase reaction suggest that the 2'-phosphate is critical for the optimum productive binding of the dinucleotide substrate. Initial velocity patterns obtained by varying 6-phosphogluconate at different fixed levels of NAD are nearly parallel with apparent competitive substrate inhibition by 6-phosphogluconate at pH 7 and below but intersect to the left of the ordinate at pH 8 and above. Dead-end inhibition studies indicate that the mechanism is random at all pH values. Data are interpreted in terms of a random mechanism with marked antagonism in the binding of NAD and 6-phosphogluconate at low pH. Deuterium isotope effects on V and V/K for either substrate are equal at pH 8, indicating that the kinetic mechanism is rapid equilibrium random. A decrease in the pH and the subsequent protonation of the active site general base or some other enzyme residue with a similar pK apparently results in the ineffective binding of NAD. The latter suggests either a link between the protonation state of this group and the conformation of the dinucleotide binding site or an interaction between the two.
Berdis
A J
AJ
Department of Biochemistry and Molecular Biology, Texas College of Osteopathic Medicine, Fort Worth 76107.
Cook
P F
PF
D002175:Candida / Q000201:enzymology* / Q000378:metabolism; D005656:Fungal Proteins / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D009249:NADP / Q000737:chemistry* / Q000378:metabolism; D010734:Phosphogluconate Dehydrogenase / Q000378:metabolism*; D013329:Structure-Activity Relationship
D005656:Fungal Proteins; D009249:NADP; D010734:Phosphogluconate Dehydrogenase