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Type II Calmodulin-Dependent Protein Kinase

II 型钙调蛋白依赖性蛋白激酶

基本信息

批准号：
9117799
负责人：
Ross Dalbey
金额：
$ 27万
依托单位：
Ohio State University Research Foundation -DO NOT USE
依托单位国家：
美国
项目类别：
Continuing Grant
财政年份：
1992
资助国家：
美国
起止时间：
1992-02-15 至 1996-01-31
项目状态：
已结题

来源：
https://www.nsf.gov/awardsearch/showAward?AWD_ID=9117799&HistoricalAwards=false
关键词：
Type II Calmodulin Dependent Protein

项目摘要

With the goal of elucidating molecular mechanisms by which calmodulin-activated enzymes aid transformation of CA2+-signals into biochemical responses, this project addresses relationships between structure, function, and regulation of the type II calmodulin-dependent protein kinase. The major target is a recombinant, monomeric form of the kinase which contains catalytic and regulatory domains but lacks the association domain. Site-specific mutagenesis will be used to produce additional forms of the kinase and calmodulin. How Mg2+, calmodulin, substrates, and autophosphorylation regulate domain folding, interactions, and topographical arrangements will be a major focus. Chemical crosslinking and active site-directed reagents will be used to probe sequences of events in kinase activation. Fluorescent probes will be attached to specific sites of calmodulin and the kinase for fluorescence and energy transfer studies; suitable metal ions and nucleotide analogs provide additional fluoroprobes. Changes in circular dichroism and in the susceptibility of the protein to proteolytic cleavage will be correlated with corresponding changes in kinase activity and calmodulin- dependence. The X-ray structure of the truncated kinase will be determined in a collaborative project. Current working models of the structure and regulation of this prominent calmodulin-dependent enzyme will be refined by computer modeling.

目的是阐明分子机制，钙调素激活酶有助于CA 2+信号的转化转化为生化反应，这个项目解决了结构、功能和调节之间的关系 II型钙调蛋白依赖性蛋白激酶。主要靶标是激酶的重组单体形式，含有催化和调节结构域，但缺乏关联域位点特异性诱变将用于产生其他形式的激酶和钙调蛋白。 Mg 2+、钙调蛋白、底物和自磷酸化调节结构域折叠、相互作用和拓扑结构安排将是一个主要焦点。化学交联和活性位点导向试剂将用于探测序列激酶激活的事件。荧光探针将被连接到钙调蛋白和激酶的特定位点，荧光和能量转移研究;合适的金属离子核苷酸类似物提供了额外的荧光探针。在圆二色性和在磁化率的变化，蛋白质到蛋白水解切割将与激酶活性和钙调蛋白的相应变化，依赖截短激酶的X射线结构将在合作项目中确定。当前工作这一突出的结构和监管模式钙调素依赖性酶将通过计算机精制建模