title: Chemical mechanism of 6-phosphogluconate dehydrogenase from Candida utilis from pH studies.
abstract: The pH dependence of kinetic parameters and dissociation constants for competitive inhibitors was determined in order to obtain information on the chemical mechanism for the 6-phosphogluconate dehydrogenase reaction from Candida utilis. A mechanism is proposed in which an active site general base accepts the proton from the 3-hydroxyl concomitant with hydride transfer at C-3; the resulting 3-keto intermediate is decarboxylated to give the enol of ribulose 5-phosphate, followed by tautomerization of the enol to the keto product with the assist of a second enzyme residue acting as a general acid. There is also a requirement for an ionized phosphate of 6-phosphogluconate and ribulose 5-phosphate for optimum binding. The maximum velocity is pH dependent, decreasing at high and low pH giving pK values of 6 and 10, while the V/K for 6-phosphogluconate decreases at low pH with a slope of 2 yielding pK values of 6.4 and 7.6, respectively, and at high pH with a slope of -1 yielding a pK of 8.2. The 6-sulfogluconate pKi profile decreases at low and high pH giving pK values of 7.1 and 8.5, respectively. The 5-phosphoarabonate and 5-phosphoarabonate pKi profiles show similar behavior giving pK values of 6.5 and 8.8, respectively, for the former and 6.8 and 8.8, respectively, for the latter. The V/K for NADP also decreases at low and high pH giving pKs of 7.5 and 8.1, while the ATP-Ribose pKi profile decreases at low and high pH giving pKs of 7.2 and 8.0.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Department of Biochemistry, Texas College of Osteopathic Medicine, Fort Worth 76107.
authors.forename: A J
authors.initials: AJ
authors.lastname: Berdis
authors.forename: P F
authors.initials: PF
authors.lastname: Cook
mesh_terms: D002175:Candida / Q000201:enzymology*; D005942:Gluconates / Q000378:metabolism / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008433:Mathematics; D008962:Models, Theoretical; D010734:Phosphogluconate Dehydrogenase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013329:Structure-Activity Relationship; D013379:Substrate Specificity
chemical_list: D005942:Gluconates; D010734:Phosphogluconate Dehydrogenase; C008884:6-phosphogluconic acid