Drug action of thapsigargin on the Ca2+ pump protein of sarcoplasmic reticulum.
Drug action of thapsigargin on the Ca2+ pump protein of sarcoplasmic reticulum.
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DOI：
10.1016/s0021-9258(18)54441-0
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发表时间：
1991-12
期刊：
影响因子：
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通讯作者：
Yoshiyuki Kijimag;E. Ogunbunmi;S. Fleischer
中科院分区：
文献类型：
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作者：
Yoshiyuki Kijimag;E. Ogunbunmi;S. Fleischer
Thapsigargin被发现是来自骨骼肌肌浆网（SR）、心脏SR和脑微粒体的细胞内Ca 2+泵蛋白的有效抑制剂。对于骨骼肌SR，毒胡萝卜素与Ca 2+泵蛋白对Ca 2+加载速率、Ca（2+）依赖性ATP酶活性和磷酸化中间体（EP）形成的完全抑制（MRc）的摩尔比约为1。用毒胡萝卜素预处理对Ca ~（2+）低亲和力的Ca ~（2+）泵蛋白（E_2态），可完全抑制ATP和Ca ~（2+）与Ca ~（2+）泵蛋白的结合。在Ca 2+（E1状态）的存在下，Ca 2+泵蛋白的失活保护毒胡萝卜素方面的Ca 2+结合和EP的形成。脑微粒体介导Ca 2+摄取到细胞内（肌醇1，4，5-三磷酸可释放）Ca 2+池中，其MRc也是化学计量的。约30%的脑微粒体的Ca 2+负载活性对毒胡萝卜素不敏感，表明存在其他Ca 2+泵系统。心脏的MRc为3.8，表明心脏SR的Ca 2+泵对毒胡萝卜素不太敏感。用蛋白激酶A磷酸化心脏SR增加了对毒胡萝卜素的敏感性，使MRc为2.8。总之，我们发现：1）毒胡萝卜素是细胞内膜（SR和内质网）Ca 2+泵蛋白的最有效抑制剂; 2）其主要抑制作用似乎优先抑制酶的E2形式; 3）心脏SR对毒胡萝卜素的敏感性低于骨骼肌SR和脑微粒体;心脏SR的蛋白激酶A治疗增强了对药物的敏感性。
Thapsigargin is found to be a potent inhibitor of the intracellular Ca2+ pump proteins from skeletal muscle sarcoplasmic reticulum (SR), cardiac SR, and brain microsomes. For skeletal muscle SR, the molar ratio of thapsigargin to Ca2+ pump protein for complete inhibition (MRc) of the Ca2+ loading rate, Ca(2+)-dependent ATPase activity, and formation of phosphorylated intermediate (EP) was approximately 1. When the Ca2+ pump protein of low affinity to Ca2+ (E2 state) was pretreated with thapsigargin, ATP and Ca2+ binding to the Ca2+ pump protein was completely inhibited. In the presence of Ca2+ (E1 state), Ca2+ pump protein was protected from inactivation by thapsigargin with respect to Ca2+ binding and EP formation. The MRc for brain microsomes, which mediate Ca2+ uptake into intracellular (inositol 1,4,5-trisphosphate-releasable) Ca2+ pools, is likewise stoichiometric. Approximately 30% of Ca2+ loading activity of brain microsomes was insensitive to thapsigargin, indicating the presence of other Ca2+ pumping system(s). The MRc for heart is 3.8, indicating that the Ca2+ pump of cardiac SR is less sensitive to thapsigargin. Phosphorylation of cardiac SR with protein kinase A increased the sensitivity to thapsigargin to MRc of 2.8. In summary, we find that: 1) thapsigargin is the most effective inhibitor of the Ca2+ pump protein of intracellular membranes (SR and endoplasmic reticulum); 2) its primary inhibitory action appears to inactivate the E2 form of the enzyme preferentially; 3) cardiac SR shows lesser sensitivity to thapsigargin than skeletal muscle SR and brain microsomes; protein kinase A treatment of cardiac SR enhances the sensitivity to the drug.