PHOTOAFFINITY LABELING OF A PROTEIN KINASE FROM BOVINE BRAIN WITH 8-AZIDOADENOSINE 3',5'-MONOPHOSPHATE
PHOTOAFFINITY LABELING OF A PROTEIN KINASE FROM BOVINE BRAIN WITH 8-AZIDOADENOSINE 3',5'-MONOPHOSPHATE
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DOI：
10.1021/bi00688a019
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发表时间：
1975-01-01
期刊：
影响因子：
2.9
通讯作者：
GREENGARD, P
中科院分区：
文献类型：
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作者：
POMERANTZ, AH;RUDOLPH, SA;GREENGARD, P
亚瑟H. Pomerantz，1 Stephen A. Rudolph，§ Boyd E. Haley，#和Paul Greengard* 摘要：8-叠氮腺苷B '。含有32 5 P的S ′-单磷酸（8-N3-CAMP）已被用作对存在于部分纯化的牛脑可溶性蛋白激酶制剂中的腺苷S ′-单磷酸（cAMP）结合位点具有特异性的光亲和标记。用标准滤纸法测定，发现8-N_3-cAMP和cAMP竞争同一结合位点。当用[32 P]-8-N3-cAMP平衡该蛋白质制备物，然后在253.7 nm下辐照时，放射性主要掺入表观分子量为49，000的蛋白质中，如十二烷基硫酸钠-聚丙烯酰胺凝胶电泳和放射自显影法所测定。这种标记的蛋白质在凝胶中与唯一被[t-32 P] ATP内源磷酸化的蛋白质共迁移，该蛋白质已被证明是蛋白激酶（H. Maeno，P. L.雷耶斯，T.上田，S. A.鲁道夫
Arthur H. Pomerantz, 1 Stephen A. Rudolph, § Boyd E. Haley,# and Paul Greengard* abstract: 8-Azidoadenosine B'. S'-monophosphate (8-N3-CAMP) containing 32 5P has been used as a photoaffinity label specific for the adenosine S'^'-monophosphate (cAMP) binding site (s) present in a partially purified prep-aration of soluble protein kinase from bovine brain. 8-N3-cAMP and cAMP were found to compete for the same binding site (s) in this preparation, as determined by a stan-dard filter assay. When this protein preparation was equilibrated with [32P]-8-N3-cAMP, and then irradiated at 253.7 nm, the incorporation of radioactivity was predominantly into a protein with an apparent molecular weight of 49,000, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography. This labeled protein comigrated in the gel with the only protein which is en-dogenously phosphorylated by [t-32P] ATP, a protein which has been shown to be the regulatory subunit of the protein kinase (H. Maeno, P. L. Reyes, T. Ueda, S. A. Rudolph,