Simultaneous analysis of NAD- and NADP-linked activities of dual nucleotide-specific dehydrogenases. Application to Leuconostoc mesenteroides glucose-6-phosphate dehydrogenase.
A method is described which enables one to assay simultaneously the NAD- and NADP-linked reactions of dehydrogenases which can utilize both coenzymes. The method is based on the fact that the thionicotinamide analogs of NADH and NADPH absorb light maximally at 400 nm, a wavelength sufficiently far removed from the absorbance maximum of NADH and NADPH to permit measurements of the simultaneous reduction of NAD+ (or NADP+) and the thionicotinamide analog of NADP+ (or NAD+). Application of the method to glucose-6-phosphate dehydrogenase from Leuconostoc mesenteroides reveals differential effects of glucose 6-phosphate concentration on the NAD- and NADP-linked reactions catalyzed by this enzyme which can not be detected by conventional assay procedures and which may have regulatory significance.
D005954:Glucosephosphate Dehydrogenase / Q000032:analysis*; D007700:Kinetics; D007934:Leuconostoc / Q000201:enzymology*; D009243:NAD; D009247:NADH, NADPH Oxidoreductases / Q000032:analysis*; D009249:NADP
D009243:NAD; D009249:NADP; D005954:Glucosephosphate Dehydrogenase; D009247:NADH, NADPH Oxidoreductases