The phosphorylation targets of p47phox, a subunit of the respiratory burst oxidase. Functions of the individual target serines as evaluated by site-directed mutagenesis.
The respiratory burst oxidase of phagocytes and B lymphocytes catalyzes the reduction of oxygen to O2- at the expense of NADPH. Dormant in resting cells, the oxidase is activated by exposing the cells to appropriate stimuli. During activation, p47phox, a cytosolic oxidase subunit, becomes extensively phosphorylated on a number of serines located between S303 and S379. To determine whether this phosphorylation is necessary for oxidase activation, we examined phorbol-elicited oxidase activity in EBV-transformed B lymphoblasts deficient in p47phox after transfection with plasmids expressing various S-->A mutants of p47phox. The mutant containing S-->A mutations involving all serines between S303 and S379 [S(303-379)A] was not phosphorylated, did not translocate to plasma membrane during activation and was almost devoid of function. As to individual serines, S379 was of special interest because (a) p47 phox S379 was phosphorylated in phorbol-activated lymphoblasts expressing wild-type p47phox, and (b) p47phox S379A failed to translocate to the membrane, and was as functionless as p47phox S(303-379)A; other single S-->A mutations had little effect on oxidase activity. These findings suggest that the phosphorylation of S379 may be important for oxidase activation in whole cells.
Faust
L R
LR
Scripps Research Institute, La Jolla, California 92037, USA.
el Benna
J
J
Babior
B M
BM
Chanock
S J
SJ
D000595:Amino Acid Sequence; D001402:B-Lymphocytes / Q000201:enzymology*; D001483:Base Sequence; D002462:Cell Membrane / Q000201:enzymology; D003600:Cytosol / Q000201:enzymology; D004789:Enzyme Activation; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009247:NADH, NADPH Oxidoreductases / Q000096:biosynthesis / Q000378:metabolism*; D009252:NADPH Dehydrogenase / Q000096:biosynthesis / Q000378:metabolism*; D019255:NADPH Oxidases; D016376:Oligonucleotides, Antisense; D010586:Phagocytes / Q000201:enzymology*; D010750:Phosphoproteins / Q000096:biosynthesis / Q000378:metabolism*; D010766:Phosphorylation; D017354:Point Mutation; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism; D012694:Serine; D014162:Transfection
D016376:Oligonucleotides, Antisense; D010750:Phosphoproteins; D011994:Recombinant Proteins; D012694:Serine; D009247:NADH, NADPH Oxidoreductases; D019255:NADPH Oxidases; C060505:neutrophil cytosolic factor 1; C022709:superoxide-forming enzyme; D009252:NADPH Dehydrogenase