Ca2+ transport in human platelet membranes. Kinetics of active transport and passive release.
Ca2+ transport in human platelet membranes. Kinetics of active transport and passive release.
复制标题
人血小板膜中的 Ca2 转运。
DOI：
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发表时间：
1986
影响因子：
4.8
通讯作者：
W. Dean
中科院分区：
文献类型：
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作者：
S. Adunyah;W. Dean
主动Ca 2+转运和被动释放的特点是在粗和纯化的人血小板膜，以便于比较与骨骼肌肌浆网。内质网标志物富集3至14倍，在纯化的膜，而表面膜抗原减少4倍，线粒体污染被完全消除。血小板膜活性Ca ~（2+）转运的最适pH值为7.6，Ca ~（2+）-ATP酶活性的最适pH值范围为7.6 ~ 8.0。加入MgATP后，活性Ca 2+转运活性出现爆发。在不存在磷酸盐的情况下，在20 s内达到稳态;添加磷酸盐促进持续摄取超过1 h。最大泵化学计量为2.0 Ca 2 +/ATP。Ca 2+离子载体A23187在磷酸盐存在下引起90%的螯合Ca 2+的快速释放。Ca 2+转运对MgATP的依赖是双相的，表观Km值为0.6 mM和9.5 μ M。动力学测量与不同的外部Ca 2+产生了一个单一的Km为0.1 μ M。Mg ~（2+）促进Ca ~（2+）转运和Ca ~（2+）-ATP酶活性。与粗品和纯化的膜的结果是相似的，并与肌浆网的钙泵比较显示几乎相同的酶性质。与比较主动钙转运的结果相反，血小板膜的钙释放特性与肌浆网的钙释放特性有很大不同。与肌浆网相反，外部Ca 2+并不促进血小板膜中螯合Ca 2+的释放。此外，ATP耗竭后，血小板膜自发释放Ca 2+并没有发生。三磷酸肌醇诱导血小板膜钙离子快速部分释放，但在相同条件下对肌浆网无影响。因此，在血小板和骨骼肌的内膜中，活性Ca 2+转运是非常相似的，但Ca 2+释放的机制似乎完全不同。
Active Ca2+ transport and passive release were characterized in crude and purified human platelet membranes to facilitate comparison with skeletal muscle sarcoplasmic reticulum. Endoplasmic reticulum markers were enriched from 3- to 14-fold in the purified membranes, while surface membrane antigens were reduced 4-fold and mitochondrial contamination was completely eliminated. The pH optimum for active Ca2+ transport in platelet membranes was 7.6, and the optimum for Ca2+-ATPase activity ranged from 7.6 to 8.0. Upon addition of MgATP there was a burst in active Ca2+ transport activity. In the absence of phosphate, steady state was reached within 20 s; added phosphate promoted continued uptake for greater than 1 h. The maximum pump stoichiometry was 2.0 Ca2+/ATP. The Ca2+ ionophore A23187 caused rapid release of 90% of the sequestered Ca2+ in the presence of phosphate. The dependence of Ca2+ transport on MgATP was biphasic with apparent Km values of 0.6 mM and 9.5 microM. Kinetic measurements with varied external Ca2+ yielded a single Km of 0.1 microM. Mg2+ stimulated Ca2+ transport and Ca2+-ATPase activities. Results with crude and purified membranes were similar, and comparison with the Ca2+ pump from sarcoplasmic reticulum revealed nearly identical enzymatic properties. In contrast to the results of comparing active Ca2+ transport, the characteristics of Ca2+ release from platelet membranes were quite different from those of sarcoplasmic reticulum. External Ca2+ did not promote release of sequestered Ca2+ from platelet membranes in contrast to sarcoplasmic reticulum. In addition, spontaneous release of Ca2+ from platelet membranes did not occur after ATP depletion. Inositol trisphosphate induced rapid partial release of Ca2+ from platelet membranes but had no effect on sarcoplasmic reticulum under identical conditions. Thus active Ca2+ transport is quite similar in internal membranes of platelet and skeletal muscle, but the mechanism of Ca2+ release appears to be entirely different.