Nagata K, Nagao S, Nozawa Y
Department of Biochemistry, Gifu University School of Medicine, Japan.
Biochem Biophys Res Commun. 1989 Apr 14;160(1):235-42. doi: 10.1016/0006-291x(89)91646-x.
We have purified and characterized two kinds of GTP-binding proteins with Mr of 22,000 in human platelet membrane (main; m22KG(I), minor; m22KG(II)) (Nagata, K. and Nozawa, Y. (1988) FEBS Lett. 238, 90-94). In this study, the main GTP-binding protein (m22KG(I)) was found to be phosphorylated by cyclic AMP-dependent protein kinase (A-kinase), but not by protein kinase C. About 0.5 mol of phosphate was maximally incorporated into one mol of the protein and this phosphorylation was inhibited in the presence of A-kinase inhibitor. Phosphorylation of m22KG(I) did not alter either its GTP-binding or GTPase activity. When m22KG(I) was incubated alone or in the presence of 100 microM guanosine 5'-(3-O-thio)triphosphate (GTP gamma S) and then exposed to A-kinase, no significant changes in the level of phosphorylation were observed. On the other hand, the most abundant GTP-binding protein with Mr of 21,000 (c21KG) in human platelet cytosol, which was identified as a transformation suppressor gene product (rap 1 protein, smg p21 and Krev-1 protein), was not phosphorylated by A-kinase under the same condition. However, c21KG was phosphorylated by A-kinase after pretreatment with alkaline phosphatase.
我们已经纯化并鉴定了人血小板膜中两种分子量为22,000的GTP结合蛋白（主要的；m22KG(I)，次要的；m22KG(II)）（永田，K.和野泽，Y.（1988年）《欧洲生物化学学会联合会快报》238，90 - 94）。在本研究中，发现主要的GTP结合蛋白（m22KG(I)）可被环磷酸腺苷依赖性蛋白激酶（A激酶）磷酸化，但不能被蛋白激酶C磷酸化。每摩尔该蛋白最多可掺入约0.5摩尔磷酸盐，并且这种磷酸化在A激酶抑制剂存在时受到抑制。m22KG(I)的磷酸化既不改变其GTP结合活性也不改变其GTP酶活性。当m22KG(I)单独孵育或在100微摩尔鸟苷5' - （3 - O - 硫代）三磷酸（GTPγS）存在下孵育，然后暴露于A激酶时，未观察到磷酸化水平有显著变化。另一方面，人血小板胞质溶胶中最丰富的分子量为21,000的GTP结合蛋白（c21KG），它被鉴定为一种转化抑制基因产物（rap 1蛋白、smg p21和Krev - 1蛋白），在相同条件下不能被A激酶磷酸化。然而，c21KG在用碱性磷酸酶预处理后可被A激酶磷酸化。
