Cell-free activation of the respiratory burst oxidase by protein kinase C.
In intact neutrophils, phorbol ester treatment activates the respiratory burst oxidase, the enzyme responsible for O2-production by phagocytes. This effect is thought to be dependent on protein kinase C and on the phosphorylation of p47phox. In this paper, we report that protein kinase C activates the respiratory burst oxidase in a cell-free system consisting of isolated neutrophil cytosol and membrane. Oxidase activation required a highly active protein kinase C, recombinant p47phow and ATP, and was inhibited by the protein kinase C inhibitors H-7 and GF-109203X. PERIl depletion of cytosolic ATP by dialysis reduced oxidase activation by over 50% In contrast, neither protein kinase C inhibitors nor ATP depletion affected oxidase activation by SDS. These findings strongly suggest that in the cell-free system, the oxidase can be activated by the phosphorylation of p47phox.
el-Benna
J
J
Department of Molecular and Experimental Medicine, Scripps Research Institute, La Jolla, CA 92037, USA.
Park
J W
JW
Ruedi
J M
JM
Babior
B M
BM
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002374:Catalase / Q000494:pharmacology; D002417:Cattle; D002474:Cell-Free System; D003600:Cytosol / Q000201:enzymology; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D007211:Indoles / Q000494:pharmacology; D007546:Isoquinolines / Q000494:pharmacology; D008301:Maleimides / Q000494:pharmacology; D009247:NADH, NADPH Oxidoreductases / Q000378:metabolism*; D009249:NADP / Q000378:metabolism; D019255:NADPH Oxidases; D009419:Nerve Tissue Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000494:pharmacology; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D010879:Piperazines / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000494:pharmacology*; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D051381:Rats; D016897:Respiratory Burst / Q000187:drug effects*; D013482:Superoxide Dismutase / Q000494:pharmacology
D004791:Enzyme Inhibitors; D007211:Indoles; D007546:Isoquinolines; D008301:Maleimides; D009419:Nerve Tissue Proteins; D010750:Phosphoproteins; D010879:Piperazines; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D009249:NADP; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D000255:Adenosine Triphosphate; D002374:Catalase; D013482:Superoxide Dismutase; D009247:NADH, NADPH Oxidoreductases; D019255:NADPH Oxidases; C060505:neutrophil cytosolic factor 1; C022709:superoxide-forming enzyme; D011493:Protein Kinase C; C070515:bisindolylmaleimide I