Petukhov S P, Grivennikov I A, Bulargina T V, Severin E S
Biokhimiia. 1984 Aug;49(8):1367-74.
The catalytic subunits of cAMP-dependent protein kinases I and II were isolated from rabbit skeletal muscles in a homogeneous state. The specific phosphotransferase activities of homogeneous preparations of catalytic subunits were 8 mumol/mg X min (type I) and 6 mumol/mg X min (type II). In order to elucidate the mechanisms of the phosphotransferase reaction, the steady-state kinetics method and an inhibitory analysis involving the phosphotransferase reaction products, ADP and phosphohistone H1, were used. It was shown that phosphorylation of histone H1 catalyzed both by protein kinases I and II occurs via a random "bi-bi" mechanism. The values of constants for kinetic equation of the phosphotransferase reaction coincide with those for the catalytic subunits of both protein kinase types and are equal to 11 microM (KmATP), 60 microM (KmH1), 5.0 microM (KSATP) and 27 microM (KSH1). The value of the competitive inhibition constant for Mg-ADP (KiADP) is also identical for the catalytic subunits of types I and II and is equal to 30 microM. In both cases, the phosphorylated histone H1 inhibits the phosphotransferase reaction; this inhibition is partly competitive with respect to histone H1.
从兔骨骼肌中以均一状态分离出环磷酸腺苷（cAMP）依赖性蛋白激酶I和II的催化亚基。催化亚基均一制剂的特异性磷酸转移酶活性分别为8 μmol/mg·min（I型）和6 μmol/mg·min（II型）。为了阐明磷酸转移酶反应的机制，采用了稳态动力学方法以及涉及磷酸转移酶反应产物二磷酸腺苷（ADP）和磷酸化组蛋白H1的抑制分析。结果表明，蛋白激酶I和II催化的组蛋白H1磷酸化均通过随机的“双底物乒乓”机制进行。磷酸转移酶反应动力学方程的常数值与两种蛋白激酶类型的催化亚基的常数值一致，分别为11 μM（ATP米氏常数KmATP）、60 μM（组蛋白H1米氏常数KmH1）、5.0 μM（ATP底物常数KSATP）和27 μM（组蛋白H1底物常数KSH1）。I型和II型催化亚基的Mg-ADP竞争性抑制常数（KiADP）值也相同，均为30 μM。在两种情况下，磷酸化的组蛋白H1均抑制磷酸转移酶反应；这种抑制对组蛋白H1而言部分具有竞争性。
