Calcium effects on calmodulin lysine reactivities.
Calcium effects on calmodulin lysine reactivities.
复制标题
钙对钙调蛋白赖氨酸反应性的影响。
DOI：
10.1016/0003-9861(87)90017-8
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发表时间：
1987
影响因子：
3.9
通讯作者：
Brew,K
中科院分区：
文献类型：
--
作者：
Giedroc,DP;Puett,D;Sinha,SK;Brew,K
个别赖氨酸对猪睾丸钙调素的差异反应性，通过高比活度[3 H]乙酸酐作为钙调素的摩尔比的函数的痕量标记来确定。在进展中从钙2 +-耗尽形式的蛋白质的钙2+：钙调蛋白的摩尔比为5：1，6的7个赖氨酰残基表现出适度的1.5至3.0倍的反应性增加。相比之下，赖氨酸75的反应性增强了20倍以上。当每个赖氨酸的反应性的变化归一化为最大变化的百分比时，发现大多数残基分为两个不同的类别。一类，包括赖氨酸94和148从两个羧基末端的钙离子结合域3和4，分别表现出约90%的反应性变化时，钙离子：钙调蛋白的摩尔比为2：1，这些残基被扰动后，进一步加入钙离子非常少。另一类，包括赖氨酸13，21，和30从氨基末端结构域1和赖氨酸75从延长螺旋连接的两个球状叶的钙调蛋白，经历了大部分的整体反应性变化（55-70%）之间的2和5当量的钙2+每摩尔的钙调蛋白。赖氨酸77是不同的，其变化模式，经历大约相等的变化与每个Ca 2+的增量。这些结果是一致的模型，其中Ca 2+首先结合到钙调蛋白的两个羧基末端位点，没有明显的偏好，伴随着未被占用的氨基末端结构域中的赖氨酸的微环境的微小改变。然后第三和第四个Ca 2+离子结合到后两个结构域，再次没有偏好的证据，在分子的羧基末端的赖氨酸反应性几乎没有变化。在中央螺旋的群体的环境似乎经历的方式，反映了他们接近的氨基和羧基末端结构域的变化。在这项工作的过程中，人们发现，赖氨酸94 apocalmodulin是特别扰动的EGTA的加入，这表明螯合剂可能与钙调蛋白或附近的第三个Ca 2+结合结构域。
The differential reactivities of individual lysines on porcine testicular calmodulin were determined by trace labeling with high specific activity [3H]acetic anhydride as a function of the molar ratio of Ca2+to calmodulin. In progressing from the Ca2+-depleted form of the protein to a Ca2+:calmodulin molar ratio of 5:1, six of the seven lysyl residues exhibited a modest 1.5- to 3.0-fold increase in reactivity. Lys 75, in contrast, was enhanced in reactivity greater than 20-fold. When the change in reactivity of each lysine was normalized as a percentage of the maximum change, most of the residues were found to fall into two distinct classes. One class, comprising lysines 94 and 148 from the two carboxy terminal Ca2+-binding domains 3 and 4, respectively, exhibited about 90% of their reactivity change when the Ca2+:calmodulin molar ratio was 2:1, and these residues were perturbed very little upon further addition of Ca2+. The other class, encompassing lysines 13, 21, and 30 from the amino terminal domain 1 and Lys 75 from the extended helix connecting the two globular lobes of calmodulin, underwent most of their overall reactivity change (55–70%) between 2 and 5 equivalents of Ca2+per mol of calmodulin. Lys 77 was distinct in its pattern of change, undergoing approximately equal changes with each Ca2+increment. These results are consistent with a model where Ca2+first binds to the two carboxy terminal sites of calmodulin with no apparent preference, concomitant with minor alterations in the microenvironments of lysines in the unoccupied amino terminal domains. The third and fourth Ca2+ions then bind to these latter two domains, again with no evidence of preference, with little change in the lysine reactivities at the carboxy terminus of the molecule. The environments of groups in the central helix appear to undergo changes in a manner that reflects their proximity to the amino and carboxy terminal domains. In the course of this work, it was found that Lys 94 in apocalmodulin is specifically perturbed by the addition of EGTA, suggesting that the chelating agent may interact with calmodulin at or near the third Ca2+-binding domain.