Neuronal NADPH diaphorase is a nitric oxide synthase
Autor: | Steven R. Vincent, Bruce T. Hope, Gregory J. Michael, Karl M. Knigge |
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Rok vydání: | 1991 |
Předmět: |
Male
Immunoblotting Fluorescent Antibody Technique Biology Nitric oxide chemistry.chemical_compound Affinity chromatography Diaphorase Animals NADPH dehydrogenase chemistry.chemical_classification Neurons Multidisciplinary Nitroblue Tetrazolium Endothelium-derived relaxing factor NADPH Dehydrogenase Brain Rats Inbred Strains Molecular biology Rats Nitric oxide synthase Molecular Weight Kinetics Enzyme chemistry Biochemistry biology.protein Amino Acid Oxidoreductases Nitric Oxide Synthase Immunostaining Research Article |
Zdroj: | Proceedings of the National Academy of Sciences of the United States of America. 88(7) |
ISSN: | 0027-8424 |
Popis: | NADPH diaphorase histochemistry selectively labels a number of discrete populations of neurons throughout the nervous system. This simple and robust technique has been used in a great many experimental and neuropathological studies; however, the function of this enzyme has remained a matter of speculation. We, therefore, undertook to characterize this enzyme biochemically. With biochemical and immunochemical assays, NADPH diaphorase was purified to apparent homogeneity from rat brain by affinity chromatography and anion-exchange HPLC. Western (immunoblot) transfer and immunostaining with an antibody specific for NADPH diaphorase labeled a single protein of 150 kDa. Nitric oxide synthase was recently shown to be a 150-kDa, NADPH-dependent enzyme in brain. It is responsible for the calcium/calmodulin-dependent synthesis of the guanylyl cyclase activator nitric oxide from L-arginine. We have found that nitric oxide synthase activity and NADPH diaphorase copurify to homogeneity and that both activities could be immunoprecipitated with an antibody recognizing neuronal NADPH diaphorase. Furthermore, nitric oxide synthase was competitively inhibited by the NADPH diaphorase substrate, nitro blue tetrazolium. Thus, neuronal NADPH diaphorase is a nitric oxide synthase, and NADPH diaphorase histochemistry, therefore, provides a specific histochemical marker for neurons producing nitric oxide. |
Databáze: | OpenAIRE |
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