Properties of the NADPH dehydrogenase component of the oxidase complex from rabbit peritoneal neutrophils: reconstitution of an oxidase activity with the dehydrogenase component and a membrane extract.
A flavin-linked NADPH cytochrome c oxido-reductase of molecular mass 77-kDa was extracted from membranes of rabbit peritoneal neutrophils and purified in the presence of Triton X-100. The redox properties of this enzyme were examined. By some criteria including its high sensitivity to mersalyl, and its relatively high specificity for NADPH compared to NADH, the rabbit neutrophil NADPH cytochrome c reductase resembled NADPH-cytochrome P-450 reductase. Limited proteolysis generated water soluble fragments, with molecular masses of 67-kDa and 57-kDa, which were still endowed with a substantial reductase activity. When added to a lysate of neutrophil membranes in octylglucoside, in the presence of an oxidase activation medium consisting of rabbit neutrophil cytosol, GTP-gamma-S, arachidonic acid and Mg2+, the purified reductase enhanced the production of O2-., suggesting that it forms part of the O2-. generating oxidase.
Laporte
F
F
LBIO/Laboratoire de Biochimie, Département de Recherche Fondamentale, Centre d'Etudes Nucléaires, Grenoble, France.
Doussiere
J
J
Vignais
P V
PV
D000818:Animals; D002248:Carbon Monoxide / Q000378:metabolism / Q000494:pharmacology; D002462:Cell Membrane / Q000201:enzymology; D004591:Electrophoresis, Polyacrylamide Gel; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006160:Guanosine Triphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D008862:Microsomes, Liver / Q000201:enzymology; D009247:NADH, NADPH Oxidoreductases / Q000378:metabolism*; D009252:NADPH Dehydrogenase / Q000378:metabolism*; D009251:NADPH-Ferrihemoprotein Reductase / Q000302:isolation & purification / Q000378:metabolism*; D009504:Neutrophils / Q000201:enzymology*; D010529:Peritoneal Cavity; D011817:Rabbits; D013053:Spectrophotometry; D013481:Superoxides / Q000378:metabolism; D013873:Thionucleotides / Q000494:pharmacology
D013873:Thionucleotides; D013481:Superoxides; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D002248:Carbon Monoxide; D006160:Guanosine Triphosphate; D009247:NADH, NADPH Oxidoreductases; D009251:NADPH-Ferrihemoprotein Reductase; D009252:NADPH Dehydrogenase; D008274:Magnesium