Deuterium isotope effects for the nonenzymatic and glutamate dehydrogenase catalyzed reduction of an α-imino acid by NADH

• Published in: Journal of the American Chemical Society 1985, Vol.107 (14), p.4301-4305
• Main Authors: SRINIVASAN, R, FISHER, H. F
• Format: Article
• Language: English
• Subjects: Biological and medical sciences; Chemistry; Delta super-pyrroline-2-carboxylic acid; deuterium; Exact sciences and technology; Fundamental and applied biological sciences. Psychology; glutamate dehydrogenase; Labelled compounds chemistry; Mechanisms. Catalysis. Electron transfer. Models; Molecular biophysics; NADPH; Organic chemistry; Physical chemistry in biology; Reactivity and mechanisms; reduction
• ISSN: 0002-7863; 1520-5126
• DOI: 10.1021/ja00300a038

The mechanisms of the nonenzymatic and glutamate dehydrogenase catalyzed reduction of an alpha -imino acid, Delta super(1)-pyrroline-2-carboxylic acid, by NAD(P)H have been studied by deuterium isotope effects. The partition isotope effects for the nonenzymatic reaction with 4-deuterated 1,4-dihydronicotinamides are about the same as the corresponding observed kinetic isotope effects with 4,4-dideuterio-1,4-dihydronicotinamides, suggesting that the hydrogen-transfer step is solely rate limiting. This reaction is characterized by an intrinsic primary kinetic isotope effect of 1.3 and a very product-like transition state. The enzymatic reaction has been studied by determining the second-order rate constants for the reduction of the imino acid by the enzyme-NADH complex with 4,4-dideuterio and stereospecifically labeled 4-deuterio NADH. The primary isotope effect when the in-place hydrogen is protium is 3.80, and the secondary isotope effect when the in-flight hydrogen is protium is 1.21. Deuteration at one site lowers the isotope effects at the other by 13%.