Andreeva L, Crompton M
Department of Biochemistry and Molecular Biology, University College London, England.
Eur J Biochem. 1994 Apr 1;221(1):261-8. doi: 10.1111/j.1432-1033.1994.tb18737.x.
Mitochondria contain a structure which forms a large aqueous pore in the inner membrane after Ca2+ overload in the presence of Pi. In the present study, pore activation in liver mitochondria was monitored using the collapse of the inner membrane potential (delta psi). Ca(2+)-induced pore opening (delta psi collapse) was prevented by the immunosuppressant cyclosporin A, but cyclosporin A did not reverse pore opening (i.e. allow delta psi regeneration) unless ADP was also added. At concentrations that produced substantial pore blockade, [3H]cyclosporin partitioned more or less equally between membrane and soluble fractions, but the distribution was shifted slightly to the membranes in the presence of ADP. ADP also increased the binding of [3H]cyclosporin A to membranes washed free of soluble components. The indication that cyclosporin A inhibition of the pore is mediated by an ADP-sensitive membrane component was examined using a tritiated photoactivable derivative of cyclosporin A. ADP selectively increased covalent binding of this derivative to a membrane component. This component eluted from molecular-sizing columns as a 13-17-kDa-protein in the presence of 0.5% Chaps as detergent and migrated as a 10-kDa (approximately) protein in SDS/PAGE. These findings provide the first evidence that a protein of approximately 10 kDa may be part of the cyclosporin-A receptor of the Ca(2+)-activated pore. The possible implications of these findings are discussed.
线粒体含有一种结构，在存在无机磷酸盐（Pi）的情况下，当Ca2+过载时，该结构会在内膜上形成一个大的水性孔道。在本研究中，利用内膜电位（δψ）的崩溃来监测肝线粒体中的孔道激活情况。免疫抑制剂环孢菌素A可阻止Ca(2+)诱导的孔道开放（δψ崩溃），但除非同时添加ADP，否则环孢菌素A不会逆转孔道开放（即允许δψ再生）。在产生显著孔道阻断的浓度下，[3H]环孢菌素在膜相和可溶性组分之间的分配大致相等，但在存在ADP的情况下，分布会略微向膜相偏移。ADP还增加了[3H]环孢菌素A与已洗涤去除可溶性成分的膜的结合。使用环孢菌素A的氚标记光活性衍生物研究了环孢菌素A对孔道的抑制作用是由ADP敏感的膜成分介导的这一迹象。ADP选择性地增加了该衍生物与膜成分的共价结合。在存在0.5% 3-[(3-胆酰胺丙基)二甲基铵]-1-丙磺酸（Chaps）作为去污剂的情况下，该成分从分子排阻柱上洗脱下来时为一种13 - 17 kDa的蛋白质，在十二烷基硫酸钠/聚丙烯酰胺凝胶电泳（SDS/PAGE）中迁移为一种约10 kDa的蛋白质。这些发现提供了首个证据，表明一种约10 kDa的蛋白质可能是Ca(2+)激活孔道的环孢菌素A受体的一部分。讨论了这些发现可能的影响。
