Phosphorylation of the respiratory burst oxidase subunit p47phox as determined by two-dimensional phosphopeptide mapping. Phosphorylation by protein kinase C, protein kinase A, and a mitogen-activated protein kinase.
Phosphorylation of the respiratory burst oxidase subunit p47phox as determined by two-dimensional phosphopeptide mapping. Phosphorylation by protein kinase C, protein kinase A, and a mitogen-activated protein kinase.
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DOI：
10.1074/jbc.271.11.6374
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发表时间：
1996-03
期刊：
影响因子：
--
通讯作者：
J. Benna;L. R. Faust;J. Johnson;B. Babior
中科院分区：
文献类型：
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作者：
J. Benna;L. R. Faust;J. Johnson;B. Babior
呼吸爆发氧化酶负责吞噬细胞和B淋巴细胞产生超氧化物（O2）。这种多组分酶在静息细胞中处于休眠状态，但在细胞暴露于适当刺激时被激活。激活后，细胞溶质氧化酶亚基p47 phox上的几个丝氨酸残基被磷酸化。利用二维胰蛋白酶磷酸肽图谱，我们研究了p47 phox在负载32 Pi的EB病毒转化的B淋巴母细胞中的磷酸化，这些淋巴母细胞表达野生型p47 phox或几种P47 phox Ser -> Ala突变体中的任何一种。我们能够鉴定野生型p47 phox的标记肽是那些含有Ser 303/304、Ser 315、Ser 320、Ser 328和/或Ser 359/370和Ser 345/348的肽，没有发现含有32 P标记的Ser 310的肽。蛋白激酶C磷酸化所有的肽，除了一个含有Ser 345/348;蛋白激酶A磷酸化的肽含有Ser 320和一个或两个含有Ser 328和Ser 359/370的肽;而促分裂原活化的蛋白激酶磷酸化的肽含有Ser 345/348。这些研究结果表明，这三种激酶在呼吸爆发氧化酶的激活中起着不同的作用，它们中的每一种都催化p47 phox中不同组丝氨酸的磷酸化。
The respiratory burst oxidase is responsible for superoxide (O2) production by phagocytes and B lymphocytes. This multicomponent enzyme is dormant in resting cells but is activated on exposure of the cells to an appropriate stimulus. Upon activation, several serine residues on the cytosolic oxidase subunit p47phox become phosphorylated. Using two-dimensional tryptic phosphopeptide mapping, we studied the phosphorylation of p47phox in 32Pi-loaded Epstein-Barr virus-transformed B lymphoblasts expressing wild type p47phox or any of several P47phox Ser -> Ala mutants. We were able to identify the labeled peptides from wild type p47phox as those contain- ing Ser303/304 Ser315, Ser320, Ser328 and/or Ser359/370, and Ser345/348 ; no 32P-labeled Ser310-containing peptide was found. When purified p47phox, was phosphorylated in vitro by various protein kinases, varying phosphopeptide patterns were observed. Protein kinase C phosphorylated all the peptides except the one containing Ser345/348; protein kinase A phosphorylated the peptide containing Ser320 and one or both of the peptides containing Ser328 and Ser359/370; while mitogen-activated protein kinase phophorylated only the peptide containing Ser345/348. These findings suggest that these three kinases play distinct roles in the activation of the respiratory burst oxidase, each of them catalyzing the phosphorylation of a different group of serines in p47phox.