title: Identification of a thermolabile component of the human neutrophil NADPH oxidase. A model for chronic granulomatous disease caused by deficiency of the p67-phox cytosolic component.
abstract: Mild heating of human neutrophils inactivates the respiratory burst oxidase, producing a defect in superoxide production and bacterial killing comparable to that seen in patients afflicted with chronic granulomatous disease (CGD). We have now investigated the mechanism and specificity of this inactivation by examining the effect of mild heating on the known oxidase components: the membrane-bound subunits of the cytochrome b558 (gp91-phox and p22-phox) and the two cytosolic oxidase factors (p47-phox and p67-phox). Heating (46 degrees C for 7.5 min) caused intact neutrophils to lose greater than 85% of their capacity to produce superoxide, a defect which was localized to the cytosolic, but not the membrane, fraction. Complementation studies with CGD cytosols deficient in either p47-phox or p67-phox suggested that the defective component of heat-inactivated cytosol was p67-phox. This was confirmed by experiments showing that recombinant p67-phox, but not p47-phox, exhibited lability at 46 degrees C and completely reconstituted oxidase activity of heat-treated cytosol. These studies indicate that mild heating of either intact neutrophils or normal neutrophil cytosol results in a selective inactivation of p67-phox, providing a model oxidase system for the extremely rare p67-phox-deficient form of CGD.
authors.affiliation: Department of Molecular and Experimental Medicine, Scripps Research Institute, La Jolla, California 92037.
authors.forename: R W
authors.initials: RW
authors.lastname: Erickson
authors.forename: S E
authors.initials: SE
authors.lastname: Malawista
authors.forename: M C
authors.initials: MC
authors.lastname: Garrett
authors.forename: G
authors.initials: G
authors.lastname: Van Blaricom
authors.forename: T L
authors.initials: TL
authors.lastname: Leto
authors.forename: J T
authors.initials: JT
authors.lastname: Curnutte
mesh_terms: D000595:Amino Acid Sequence; D002462:Cell Membrane / Q000201:enzymology; D003573:Cytochrome b Group / Q000737:chemistry* / Q000172:deficiency / Q000378:metabolism; D003600:Cytosol / Q000201:enzymology; D005816:Genetic Complementation Test; D006105:Granulomatous Disease, Chronic / Q000201:enzymology*; D006358:Hot Temperature; D006801:Humans; D008969:Molecular Sequence Data; D009247:NADH, NADPH Oxidoreductases / Q000737:chemistry*; D019255:NADPH Oxidases; D009504:Neutrophils / Q000201:enzymology*; D016897:Respiratory Burst; D013481:Superoxides / Q000378:metabolism
chemical_list: D003573:Cytochrome b Group; D013481:Superoxides; C036888:cytochrome b558; D009247:NADH, NADPH Oxidoreductases; D019255:NADPH Oxidases