Structure and Function of Calmodulin-dependent Protein Kinase II and Its Physiological Significance

通过对大鼠脑提取液的cDNA序列分析，发现了四种不同的钙调蛋白依赖蛋白激酶II(CaM-Kinase II)多肽(亚型)，即α、β、γ和Delta，它们来自四个不同的基因。用针对不同mRNAs的探针对不同组织的提取物进行RNA印迹杂交分析表明，γ和βmRNAs在不同的组织中表达，而α和βmRNAs主要在大脑中表达，CaM-Kinase II的自磷酸化被认为是控制其蛋白激酶活性的重要自我调节机制。即使在有钙调素存在的情况下，没有发生自动磷酸化的酶也只表现出最大活性的15%，但将该酶暴露在钙调素中会导致该酶在Thr^&lt；286&gt；上快速自动磷酸化，从而将酶转化为最具活性的形式。活性形式即使在…中也具有蛋白激酶活性更多的是没有钙调素，尽管在没有钙离子的情况下对蛋白质底物的亲和力要低得多；酪氨酸羟基酶催化儿茶酚胺生物合成中的限速步骤。已知的是，催化儿茶酚胺生物合成的限速步骤的酪氨酸羟基酶被三种第二信使反应的多功能蛋白激酶所磷酸化，这三种多功能蛋白激酶是cAMP依赖的蛋白激酶(A-Kinase)、蛋白激酶C(C-Kinase)和CaM-Kinase II。然而，该酶通过A-Kinase的磷酸化而被激活，而不被C-Kinase激活，只有在激活蛋白存在的情况下才被CaM-Kinase II激活。我们推测了这三种多功能蛋白激酶对酶活性的可能调节机制。另外一种几乎只在大脑中存在的钙调蛋白依赖的蛋白激酶，钙调蛋白依赖的蛋白激酶IV(CaM-Kinase IV)具有广泛的底物特异性，提示其在钙调素调节脑功能中的生理意义；有趣的是，CaM-Kinase IV通过自身磷酸化被激活，但通过A-Kinase的磷酸化而失活。较少

Four different polypeptides (isoforms) of calmodulin-dependent protein kinase II (CaM-kinase II), alpha, beta, gamma, and delta, which derived from four different genes, were revealed by cDNA sequence analysis of the rat brain extract. RNA blot bybridization analysis of the extracts from various tissues with probes specific for the respective mRNAs showed that gamma and delta mRNAs were expressed in various tissues, while alpha and beta mRNAs were primarily, if not exclusively, expressed in brain.Autophosphorylation of CaM-kinase II is thought to be an important selfregulatory mechanism for controlling its protein kinase activity. The enzyme that had not undergone autophosphorylation exhibited only 15% of the maximum activity even in the presence of Ca^<2+>/calmodulin, but exposure of the enzyme to Ca^<2+>/calmodulin caused rapid autophosphorylation of the enzyme on Thr^<286>, thereby converting the enzyme to the most active form. The active form had the protein kinase activity even in … More the absence of Ca^<2+>/calmodulin, although the affinity for the protein substrates were much lower in the absence of Ca^<2+> than in its presence.Tyrosine hydroxylase catalyzing the rate-limiting step in the biosynthesis of catecholamines is known to be phosphorylated by three second messenger-responsive multifunctional protein kinases, cAMP-dependent protein kinase (A-kinase), protein kinase C (C-kinase), and CaM-kinase II. However, the enzyme was activated via phosphorylation by A-kinase, not acivated by C-kinase, and activated only in the presence of activator protein by CaM-kinase II. The possible regulatory mechanism of the enzyme activity by these three multifunctional protein kinases were suggested.Another calmodulin-dependent protein kinase occurring almost exclusively in brain, calmodulin-dependent protein kinase IV (CaM-kinase IV), was found to show a broad substrate specificity, suggesting its physiological significance in the regulation of the brain function by Ca^<2+>.Interestingly, CaM-kinase IV was activated by autophosphorylation but inactivated via phosphorylation by A-kinase. Less