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Annexin A6 at the cardiac myocyte sarcolemma--evidence for self-association and binding to actin.

机译:心肌肌肉瘤处的膜联蛋白A6-自我结合和与肌动蛋白结合的证据。

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摘要

The plasma membrane of the heart muscle cell and its underlying cytoskeleton are vitally important to the function of the heart. Annexin A6 is a major cellular calcium and phospholipid binding protein. Here we show that annexin A6 copurifies with sarcolemma isolated from pig heart. Two pools of annexin A6 are present in the sarcolemma fraction, one dependent on calcium and one that resists extraction by the calcium chelator EGTA. Potential annexin A6 binding proteins in the sarcolemma fraction were identified using Far Western blotting. Two major annexin A6 binding proteins were identified as actin and annexin A6 itself. Annexin A6 bound to itself both in the presence and in the absence of calcium ions. Sites for self association were mapped by performing Western blots on proteolytic fragments of recombinant annexin A6. Annexin A6 bound preferentially not only to the N terminal fragment (domains I-IV, residues 1-352) but also to C-terminal fragments corresponding to domains V+VI and domains VII+VIII. Actin binding to annexin A6 was calcium-dependent and exclusively to the N-terminal fragment of annexin A6. A calcium-dependent complex of annexin A6 and actin may stabilize the cardiomyocyte sarcolemma during cell stimulation.
机译:心肌细胞的质膜及其潜在的细胞骨架对于心脏的功能至关重要。膜联蛋白A6是主要的细胞钙和磷脂结合蛋白。在这里,我们显示膜联蛋白A6与从猪心脏分离的肌膜共纯化。肌膜部分中存在两个膜联蛋白A6池,一个池依赖钙,另一个池抵抗钙螯合剂EGTA的提取。使用Far Western印迹法鉴定肌膜部分中潜在的膜联蛋白A6结合蛋白。两种主要的膜联蛋白A6结合蛋白被鉴定为肌动蛋白和膜联蛋白A6本身。在存在和不存在钙离子的情况下,膜联蛋白A6均与自身结合。通过在重组膜联蛋白A6的蛋白水解片段上进行Western印迹来定位自我结合的位点。膜联蛋白A6不仅优先结合N末端片段(结构域I-IV,残基1-352),而且优先结合对应于结构域V + VI和结构域VII + VIII的C末端片段。肌动蛋白与膜联蛋白A6的结合是钙依赖性的,并且仅与膜联蛋白A6的N端片段结合。膜联蛋白A6和肌动蛋白的钙依赖性复合物可以在细胞刺激过程中稳定心肌肉瘤。

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