title: Purification and characterization of glucose-6-phosphate dehydrogenase from Cryptococcus neoformans: identification as "nothing dehydrogenase".
abstract: Glucose-6-phosphate dehydrogenase (EC 1.1.1.49) was purified from Cryptococcus neoformans, a basidiomyceteous yeast that is an opportunistic pathogen of AIDS patients. The enzyme had a subunit molecular weight of 5 x 10(4), a specific activity of 50 units mg-1, and Km values for NADP and glucose-6-phosphate of 1.6 and 24 microM, respectively. The enzyme catalyzed the dehydrogenation of glucose, in the presence of dimethylsulfoxide, with Km of 5 mM and Vmax 10% of that with glucose-6-phosphate. pH profiles indicated the presence of a group with pKa of 6.6 that is involved in catalysis, and groups with pKas of about 8.8 that are involved in binding of NADP and glucose-6-phosphate. The enzyme was inhibited by NADPH, competitive versus NADP, with Ki of 1 microM, and by zinc ion, competitive versus glucose-6-phosphate, with Ki of 2 microM. Crude enzyme extract catalyzed an appreciable rate of reduction of NADP in the absence of added substrate, a "nothing dehydrogenase" activity. This activity was shown to be due to the presence of glucose-6-phosphate in the crude extract. It was calculated that cells of C. neoformans contain about 25 mumol of glucose-6-phosphate per gram, wet weight.
authors.affiliation: Department of Biochemistry and Anaerobic Microbiology, Virginia Polytechnic Institute and State University, Blacksburg 24061-0308.
authors.forename: W G
authors.initials: WG
authors.lastname: Niehaus
authors.forename: T C
authors.initials: TC
authors.lastname: Mallett
mesh_terms: D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D003455:Cryptococcus neoformans / Q000201:enzymology*; D005954:Glucosephosphate Dehydrogenase / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D046911:Macromolecular Substances; D008970:Molecular Weight; D009249:NADP / Q000378:metabolism / Q000494:pharmacology; D010084:Oxidation-Reduction; D013379:Substrate Specificity; D015032:Zinc / Q000494:pharmacology
chemical_list: D046911:Macromolecular Substances; D009249:NADP; D005954:Glucosephosphate Dehydrogenase; D015032:Zinc