Photodependent inhibition of rat liver NAD(P)H:quinone acceptor oxidoreductase by (A)-2-azido-NAD+ and (A)-8-azido-NAD |
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Authors: | P S Deng S H Zhao T Iyanagi S A Chen |
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Institution: | Division of Immunology, Beckman Research Institute of the City of Hope, Duarte, California 91010. |
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Abstract: | Two photoaffinity analogues of NAD+, (A)-2-azido-NAD+ nicotinamide 2-azidoadenine dinucleotide] and (A)-8-azido-NAD+ nicotinamide 8-azidoadenine dinucleotide], have been synthesized, and their reactivities with the rat liver NAD(P)H:quinone acceptor oxidoreductase have been investigated. The reduce nicotinamide nucleotide probes, (A)-2-azido-NADH and (A)-8-azido-NADH, were shown to be substrates of the quinone reductase. This enzyme was inhibited by (A)-8-azido-NADH, were shown to be substrates of the quinone reductase. This enzyme was inhibited by (A)-2-azido-NAD+ and (A)-8-azido-NAD+ in a photodependent manner, and the inhibition of the enzyme could be prevented by the presence of nicotinamide nucleotide substrates during photolysis. (A)-2-Azido-NAD+ was demonstrated to be a more potent inhibitor than (A)-8-azido-NAD+. In addition, the photodependent inhibition by (A)-8-azido-NAD+ increased when menadione, the substrate of the enzyme, was present during the photolysis, while menadione protected the enzyme from the photodependent inhibition by (A)-2-azido-NAD+. These results indicate that these two NAD+ analogues can be used to identify the nicotinamide nucleotide binding site of this quinone reductase and that they probably bind to the enzyme in different fashions. |
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