A one-step PMR determination of hydrogen transfer stereospecificity of NADP+-linked oxidoreductases.
1. A simple, facile one-step method has been devised to measure the stereospecificity of NADP+-linked oxidoreductases. The procedure involves coupling the test enzymes to enzymes of known stereospecificity in the presence of deuterated substrates. The regenerated NADP+ in the coupled reactions is analyzed by PMR for its deuterium content at the carbon-4 position of the nicotinamide ring. 2. It is found that malate dehydrogenase (EC 1.1.1.37), lactate dehydrogenase (EC 1.1.1.27) and glycerate dehydrogenase (EC 1.1.1.29) are A-side stereospecific whereas glutamate dehydrogenase (EC 1.4.1.3) and glycerol-3-phosphate dehydrogenase (EC 1.1.1.8) are B-side stereospecific. 3. Enzymes which can utilize both NAD+ and NADP+ have the same stereospecificity with respect to the coenzyme.
D000818:Animals; D002417:Cattle; D003067:Coenzymes / Q000378:metabolism; D006859:Hydrogen / Q000032:analysis*; D009682:Magnetic Resonance Spectroscopy; D009247:NADH, NADPH Oxidoreductases / Q000378:metabolism*; D011817:Rabbits; D013237:Stereoisomerism; D013552:Swine
D003067:Coenzymes; D006859:Hydrogen; D009247:NADH, NADPH Oxidoreductases