Wombacher H, Reuter-Smerdka M, Körber F
Curr Probl Clin Biochem. 1976;6:114-22.
Rat kidney plasma membranes prepared by the method of FITZPATRICK et al. (J Biol. Chem. 244, 3561, 1969) show protein kinase activity as well as specific cAMP binding activity (diss. const. 1.3 x 10-9 M). However, no stimulation of kinase activity by cAMP is observed in presence of exogenous substrates (e.g. histone) and only poor stimulation with endogenous substrates in the membrane could be shown. At high ionic strength (1 M NaCl) cAMP independent protein kinase activity can be solubilized. Low ionic strength buffer (1 mM Tris-HCl pH 7.4 1 mM EDTA) and non-ionic detergents (Lubrol PX, Lubrol WX and Triton X 100) are able to solubilize both protein kinase activity and cAMP binding activity. Protein kinase activity seemed to be only loosely associated with the membrane, whereas cAMP binding protein appears to be more firmly fixed into the membrane structure. In addition we have found that membranes serve as a good substrate for cytosol protein kinase (s) and Ca-ion concentration influences the effect of cAMP on protein kinase activity. Dependent on the increase of Ca-ion concentration the effect of cAMP on protein kinase changes from activation to inhibition.
用菲茨帕特里克等人（《生物化学杂志》244卷，3561页，1969年）的方法制备的大鼠肾质膜显示出蛋白激酶活性以及特异性的环磷酸腺苷（cAMP）结合活性（解离常数为1.3×10⁻⁹M）。然而，在外源底物（如组蛋白）存在的情况下，未观察到cAMP对激酶活性的刺激作用，并且仅能显示出膜中内源性底物对激酶活性的微弱刺激。在高离子强度（1M氯化钠）下，可溶解不依赖cAMP的蛋白激酶活性。低离子强度缓冲液（1mM Tris-HCl，pH 7.4，1mM乙二胺四乙酸）和非离子去污剂（Lubrol PX、Lubrol WX和Triton X 100）能够溶解蛋白激酶活性和cAMP结合活性。蛋白激酶活性似乎仅与膜松散结合，而cAMP结合蛋白似乎更牢固地固定在膜结构中。此外，我们发现质膜是胞质溶胶蛋白激酶的良好底物，并且钙离子浓度会影响cAMP对蛋白激酶活性的作用。随着钙离子浓度的增加，cAMP对蛋白激酶的作用从激活转变为抑制。
