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Multiple conformations of physiological membrane-bound cytochrome c

机译：生理膜结合细胞色素c的多种构型

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One-tenth of cytochrome c (cyt c) remains bound to the inner mitochondrial membrane (IMM) at physiological ionic strength (I; i.e., I approximate to 150 mM), exhibiting decreased electron transport (ET) activity. We now show that this form of membrane-bound cyt c (MB-cyt c) can be obtained in vitro and that binding to membranes at low I generates an additional conformation with higher ET activity, This low I bound form of MB-cyt c (MBL-cyt c) exhibited intrinsic ET rates similar to those of electrostatically bound cyt c (EB-cyt c). The ET activity of IMM-bound MB-cyt c approached slowly that of MBL-cyt c or EB-cyt c, suggesting that MB-cyt c converts to MBL-cyt c while bound to IMM, When maintained at physiological I, both forms of MB-cyt c were released from the membrane, indicating that they convert to an EB-cyt c-like form. This process may be very dynamic in cellular mitochondria, as binding and release for both MB-cyt c forms increased considerably with temperature. I-Dependent binding of MB-cyt c does not require IMM, and it can be reproduced using large or small unilamellar vesicles (SUV). Using SUV-cyt c complexes, we characterized the secondary structure of MB-cyt c and MBL-cyt c by circular dichroism. Conformational analysis revealed that cyt c binding as MB-cyt c decreases its alpha-helical content (70-79%) and increases its beta-sheet up to 135%. The secondary structure of MBL-cyt c was similar to that of EB-cyt c and soluble cyt c, with a modest increase in beta-sheet, Taken together, our experiments suggest that physiological cyt c exists in soluble and membrane-bound conformations with similar ET activity, which may exchange very rapidly, and that soluble hydrophilic proteins can bind transiently to biomembranes. [References: 65]
机译：十分之一的细胞色素c（cyt c）仍以生理离子强度（I;即I约为150 mM）与线粒体内膜（IMM）结合，表现出降低的电子传输（ET）活性。我们现在表明，这种形式的膜结合的cyt c（MB-cyt c）可以在体外获得，并且在低I时与膜的结合产生具有更高ET活性的其他构象。这种低I结合的MB-cyt c （MBL-cyt c）的内在ET速率与静电结合的cyt c（EB-cyt c）相似。与IMM结合的MB-cyt c的ET活性接近于MBL-cyt c或EB-cyt c的ET活性，这表明MB-cyt c在与IMM结合时会转化为MBL-cyt c。 MB-cyt c的一部分从膜中释放出来，表明它们转化为EB-cyt c样形式。该过程在细胞线粒体中可能是非常动态的，因为两种MB-cyt c形式的结合和释放都随温度而显着增加。 MB cyt c的I依赖性结合不需要IMM，并且可以使用大或小的单层囊泡（SUV）进行复制。使用SUV-cyt c配合物，我们通过圆二色性表征了MB-cyt c和MBL-cyt c的二级结构。构象分析显示，作为MB-cyt c的cyt c结合会降低其α-螺旋含量（70-79％），并将其β-折叠层增加至135％。 MBL-cyt c的二级结构与EB-cyt c和可溶性cyt c相似，但β-sheet有所增加。综上所述，我们的实验表明生理性cyt c以可溶性和膜结合构象存在。具有类似的ET活性，可以非常迅速地交换，并且可溶性亲水蛋白可以与生物膜瞬时结合。 [参考：65]

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