Purification and characterization of hepatic microsomal NADPH cytochrome c reductase from rhesus monkey (Macaca mulatta).
Hepatic microsomal NADPH cytochrome c reductase from nonhuman primate (Macaca mulatta) has been purified 76 fold by the combination of anion exchange, affinity and molecular sieve chromatographies. The purified preparation had approximate molecular wt of 63 kDa and carried out NADPH oxidation, cytochrome c reduction, 2,6-dichlorophenol indophenol (DCIP) reduction and production of superoxide anions (O2-.). The enhancement in NADPH cytochrome c reductase catalyzed NADPH oxidation by metal chelators viz. ethylenediaminetetra-acetic acid (EDTA)-FeCl3 and diethylenetriaminepenta-acetic acid (DTPA)-FeCl3 was dramatically higher than the enhancement in the reduction of cytochrome c and DCIP. DTPA-FeCl3 was found to be more potent stimulator of NADPH oxidation as compared to EDTA-FeCl3, but both had similar potency as for reduction of cytochrome c and DCIP were concerned. Superoxide dismutase (SOD) decreased EDTA-FeCl3 enhanced reduction of cytochrome c by 15%, but had no effect on the NADPH oxidation and DCIP reduction, whereas it significantly enhanced DTPA-FeCl3 stimulated NADPH oxidation, decreased cytochrome c reduction by 8% and did not affect DCIP reduction. In addition, SOD almost completely blocked the NADPH cytochrome c reductase catalyzed superoxide anion production. The results demonstrate that like rodents and lagomorphs, the hepatic microsomal NADPH cytochrome c reductase in nonhuman primate, Macaca mulatta can carry out single electron reduction of molecular oxygen.
Department of Biochemistry, Postgraduate Institute of Medical Education and Research, Chandigarh, India.
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D000818:Animals; D002733:Chlorophenols / Q000378:metabolism; D002846:Chromatography, Affinity; D002852:Chromatography, Ion Exchange; D003574:Cytochrome c Group / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D007502:Iron Chelating Agents / Q000494:pharmacology; D008253:Macaca mulatta; D008297:Male; D008862:Microsomes, Liver / Q000187:drug effects / Q000201:enzymology*; D008970:Molecular Weight; D009249:NADP / Q000378:metabolism; D009251:NADPH-Ferrihemoprotein Reductase / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D010084:Oxidation-Reduction / Q000187:drug effects; D013482:Superoxide Dismutase / Q000494:pharmacology
D002733:Chlorophenols; D003574:Cytochrome c Group; D007502:Iron Chelating Agents; D009249:NADP; D013482:Superoxide Dismutase; D009251:NADPH-Ferrihemoprotein Reductase; C032729:2,6-dichlorophenol