Kelly P T, Shields S, Conway K, Yip R, Burgin K
Department of Neurobiology and Anatomy, University of Texas Health Science Center, Houston 77225.
J Neurochem. 1987 Dec;49(6):1927-40. doi: 10.1111/j.1471-4159.1987.tb02456.x.
Synaptic junctions (SJs) from rat forebrain were isolated at increasing postnatal ages and examined for endogenous protein kinase activities. Our studies focused on the postnatal maturation of the multifunctional protein kinase designated Ca2+/calmodulin-dependent protein kinase II (CaM-kinase II). This kinase is comprised of a major 50-kilodalton (kDa) and a minor 60-kDa subunit. Experiments examined the developmental properties of CaM-kinase II associated with synaptic plasma membranes (SPMs) and synaptic junctions (SJs), as well as the holoenzyme purified from cytosolic extracts. Large developmental increases in CaM-kinase II activity of SJ fractions were observed between postnatal days 6 and 20; developmental changes were examined for a number of properties including (a) autophosphorylation, (b) endogenous substrate phosphorylation, (c) exogenous substrate phosphorylation, and (d) immunoreactivity. Results demonstrated that forebrain CaM-kinase II undergoes a striking age-dependent change in subunit composition. In early postnatal forebrain the 60-kDa subunit constitutes the major catalytic and immunoreactive subunit of the holoenzyme. The major peak of CaM-kinase II activity in SJ fractions occurred at approximately postnatal day 20, a time near the end of the most active period of in vivo synapse formation. Following this developmental age, CaM-kinase II continued to accumulate at SJs; however, its activity was not as highly activated by Ca2+ plus calmodulin.
在出生后不同年龄阶段分离大鼠前脑的突触连接（SJ），并检测其内源性蛋白激酶活性。我们的研究聚焦于多功能蛋白激酶——钙/钙调蛋白依赖性蛋白激酶II（CaM-激酶II）的出生后成熟过程。该激酶由一个主要的50千道尔顿（kDa）亚基和一个次要的60-kDa亚基组成。实验检测了与突触质膜（SPM）和突触连接（SJ）相关的CaM-激酶II的发育特性，以及从胞质提取物中纯化的全酶的发育特性。在出生后第6天至第20天之间，观察到SJ组分中CaM-激酶II活性有显著的发育性增加；对包括（a）自身磷酸化、（b）内源性底物磷酸化、（c）外源性底物磷酸化和（d）免疫反应性在内的一些特性进行了发育变化检测。结果表明，前脑CaM-激酶II的亚基组成发生了显著的年龄依赖性变化。在出生后早期前脑中，60-kDa亚基构成了全酶的主要催化和免疫反应性亚基。SJ组分中CaM-激酶II活性的主要峰值出现在出生后约第20天，这一时期接近体内突触形成最活跃期的末期。在这个发育年龄之后，CaM-激酶II继续在SJ处积累；然而，其活性不再像由钙离子加钙调蛋白高度激活。
