Regulation of the Function of CNS by Ca^<2+>/calmodulin-dependent Multifunctional Protein Kinases

Ca^2/钙调蛋白依赖性多功能蛋白激酶对中枢神经系统功能的调节

基本信息

批准号：
04404024
负责人：
FUJISAWA Hitoshi
金额：
$ 14.08万
依托单位：
Asahikawa Medical College
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (A)
财政年份：
1992
资助国家：
日本
起止时间：
1992 至 1994
项目状态：
已结题

项目摘要

Ca^<2+>/calmodulin-dependent protein kinase II (CaM-kinase II) and IV (CaM-kinase IV), which exist particularly abundantly in the brain, are second messenger-responsive multifunctional protein kinases, just like cAMP-dependent protein kinase and protein kinase C,and therefore these protein kinases are thought to play important roles in signal transduction in the central nervous system (CNS). The present study focuses our attention on regulatory mechanisms of the activities of CaM-kinase II and CaM-kinase IV.The autoinhibitory domain of CaM-kinase II,which is known to keep the enzymeless active in the absence of Ca^<2+>/calmodulin, was found to strongly stabilize the enzyme. Thus, CaM-kinase II appears to be in an inactive and stable state in a cell when its activity is not necessary. Ca^<2+>/calmodulin required for the activity of CaM-kinase II,on the other hand, inactivated the enzyme. Such two opposite effects of calmodulin were also observed with CaM-kinase IV,although the inactivation of CaM-kinase IV by Ca^<2+>/calmodulin was reversible in contrast to the case of CaM-kinase II.Detailed sequence analysis of CaM-kinase II cDNA clones from the brain and liver suggested a possible mechanism for producing many CaM-kinase II isoforms from the alpha, beta, gamma, and delta genes of CaM-kinase II by alternative splicing.The most prominent result of our study is the discovery of CaM-kinase IV kinase in the brain which activates CaM-kinase IV 30- to 40-fold by phosphorylation. CaM-kinase IV kinase was purified from rat cerebral cortex to apparent homogeneity upon SDS-polyacrylamide gel electrophoresis, and its molecular and catalytic properties were examined. CaM-kinase IV kinase was a Ca^<2+>/calmodulin-dependent protein kinase which phosphorylated CaM-kinase IV specifically. Human CaM-kinase IV cDNA was cloned from a cDNA library of a Jurkat cell, a cell line of human T-cell, and its nucleotide sequence and the deduced amino acid sequence were determined.

Ca^<2+>/依赖钙调蛋白的蛋白激酶II（凸轮激酶II）和IV（Cam-激酶IV），在大脑中特别丰富存在，是第二个允许的多功能蛋白激酶，就像cAMP依赖性蛋白质激酶和蛋白质激酶Cin酶和这些蛋白质酶相似的蛋白质酶（这些蛋白质酶）中的启动（这些蛋白质酶）中的启动（这些蛋白质酶）中的启动（这些蛋白质酶）中的启示性（类似于这些蛋白质酶），因此中心启动了这些含量。本研究将我们的注意力集中在Cam-激酶II和Cam-激酶IV活性的调节机制上。CAM-激酶II的自抑制结构域，在没有CA^<2+>/钙调蛋白的情况下，发现该酶使酶保持活跃，从而强烈稳定酶。因此，当不需要活性时，凸轮激酶II在细胞中似乎处于不活跃和稳定状态。另一方面，CA^<2+>/CAM-激酶II活性所需的钙调蛋白使酶失活。尽管CA^<2+>/CA^<2+>/钙调蛋白与凸轮激酶II. cam-激酶II. cam-激酶II的序列分析相比，ca^<2+>/钙调蛋白的cam-激酶IV灭活是cam-激酶II的cam-激酶II cDNA cDNA cDNA CLONE的cam-激酶II可能会产生许多可能的机动化，但与cam-激酶II分析相比，ca^<2+>/钙调蛋白与cam-激酶II分析相比，ca^<2+>/钙调蛋白与cam-激酶II相反，这可能会产生许多可能的机动体，这与ca-^<2+>/ca-ii。通过替代剪接的凸轮激酶II的三角洲基因。我们研究的最突出的结果是在大脑中发现凸轮激酶IV激酶，通过磷酸化激活30至40倍的Cam-激酶IV。 CAM-激酶IV激酶从大鼠脑皮质纯化为SDS-聚丙烯酰胺凝胶电泳的明显同质性，并检查了其分子和催化特性。 CAM-激酶IV激酶是CA^<2+>/钙调蛋白依赖性蛋白激酶，该蛋白激酶磷酸化cam-激酶IV。从Jurkat细胞的cDNA文库，人类T细胞的细胞系，其核苷酸序列和推导的氨基酸序列从Jurkat细胞的cDNA文库中克隆人。