| Abstract: | The purified mitochondrial NADH dehydrogenase enzyme has been shown to catalyze a rapid 4B-3H] NADH-H2O exchange reaction. When the enzyme is subjected to a single freeze-thaw cycle there is a complete loss of NADH dehydrogenation without a measurable decrease in the 4B-3H] NADH-H2O exchange. Complete loss of the 4B-3H] NADH-H2O exchange follows brief exposure to ultraviolet photoirradiation. The differential sensitivity of the water exchange reaction and the dehydrogenase activity suggests a direct involvement of the enzymes flavin cofactor in the catalysis of the 4B-3H] NADH-H2O exchange. Arylazido-beta-alanyl NAD+ (A3'-0-3-N-4-azido-2-nitrophenyl)amino] propionyl]NAD+) is shown to be a potent photodependent inhibitor of the 4B-3H] NADH-H2O exchange activity following photoirradiation with visible light. This is consistent with the observed photodependent inhibition of the dehydrogenase activity by this photoprobe (Chen, S. and Guillory, R.J. (1981) J. Biol. Chem. 256, 8318-8323). |
