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首页> 外文期刊>Proceedings of the National Academy of Sciences of the United States of America >Ca~(2+)-dependent structural rearrangements within Na~+-Ca~(2+) exchanger dimers
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Ca~(2+)-dependent structural rearrangements within Na~+-Ca~(2+) exchanger dimers

机译:Na〜+ -Ca〜(2+)交换二聚体中Ca〜(2+)依赖的结构重排

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摘要

Cytoplasmic Ca~(2+) is known to regulate Na~+-Ca~(2+) exchanger (NCX) activity by binding to two adjacent Ca~(2+)-binding domains (CBD1 and CBD2) located in the large intracellular loop between trans-membrane segments 5 and 6. We investigated Ca~(2+)-dependent movements as changes in FRET between exchanger proteins tagged with CFP or YFP at position 266 within the large cytoplasmic loop. Data indicate that the exchanger assembles as a dimer in the plasma membrane. Addition of Ca~(2+) decreases the distance between the cytoplasmic loops of NCX pairs. The Ca~(2+)-dependent movements detected between paired NCXs were abolished by mutating the Ca~(2+) coordination sites in CBD1 (D421A, E451A, and D500V), whereas disruption of the primary Ca~(2+) coordination site in CBD2 (E516L) had no effect. Thus, the Ca~(2+)-induced conforma-tional changes of NCX dimers arise from the movement of CBD1. FRET studies of CBD1, CBD2, and CBD1-CBD2 peptides displayed Ca~(2+)-dependent movements with different apparent affinities. CBD1-CBD2 showed a Ca~(2+)-dependent phenotype mirroring full-length NCX but distinct from both CBD1 and CBD2.
机译:已知细胞质Ca〜(2+)通过与位于大细胞内的两个相邻Ca〜(2+)结合域(CBD1和CBD2)结合来调节Na〜+ -Ca〜(2+)交换子(NCX)的活性跨膜段5和6之间的环。我们研究了Ca〜(2+)依赖的运动,即在大细胞质环中位置266处用CFP或YFP标记的交换蛋白之间FRET的变化。数据表明交换器在质膜中组装成二聚体。 Ca〜(2+)的添加减少了NCX对的胞质环之间的距离。通过突变CBD1(D421A,E451A和D500V)中的Ca〜(2+)配位点而取消了配对NCX之间检测到的Ca〜(2+)依赖性运动,而主要Ca〜(2+)配位的破坏CBD2(E516L)中的位点无效。因此,Ca〜(2+)诱导的NCX二聚体的构象变化是由CBD1的运动引起的。对CBD1,CBD2和CBD1-CBD2肽的FRET研究表明,Ca〜(2+)依赖性运动具有不同的表观亲和力。 CBD1-CBD2表现出Ca〜(2+)依赖性表型,反映了全长NCX,但不同于CBD1和CBD2。

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    Departments of Medicine (Cardiology), David Geffen School of Medicine, University of California, Los Angeles, CA 90095-1751 Departments of Cardiovascular Research Laboratory, David Geffen School of Medicine, University of California, Los Angeles, CA 90095-1751;

    Departments of Cardiovascular Research Laboratory, David Geffen School of Medicine, University of California, Los Angeles, CA 90095-1751 Departments of Physiology , David Geffen School of Medicine, University of California, Los Angeles, CA 90095-1751;

    Departments of Medicine (Cardiology), David Geffen School of Medicine, University of California, Los Angeles, CA 90095-1751 Departments of Cardiovascular Research Laboratory, David Geffen School of Medicine, University of California, Los Angeles, CA 90095-1751 Departments of Physiology , David Geffen School of Medicine, University of California, Los Angeles, CA 90095-1751;

    Departments of Medicine (Cardiology), David Geffen School of Medicine, University of California, Los Angeles, CA 90095-1751 Departments of Cardiovascular Research Laboratory, David Geffen School of Medicine, University of California, Los Angeles, CA 90095-1751 Departments of Physiology , David Geffen School of Medicine, University of California, Los Angeles, CA 90095-1751;

    Departments of Cardiovascular Research Laboratory, David Geffen School of Medicine, University of California, Los Angeles, CA 90095-1751 Departments of Physiology , David Geffen School of Medicine, University of California, Los Angeles, CA 90095-1751;

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