From the Department of Chemistry, University of California, Davis, California 95616.
the Department of Physiology and Biophysics, University of Washington, Seattle, Washington 98195, and.
J Biol Chem. 2014 Nov 7;289(45):31262-73. doi: 10.1074/jbc.M114.604439. Epub 2014 Sep 25.
CaBP4 modulates Ca(2+)-dependent activity of L-type voltage-gated Ca(2+) channels (Cav1.4) in retinal photoreceptor cells. Mg(2+) binds to the first and third EF-hands (EF1 and EF3), and Ca(2+) binds to EF1, EF3, and EF4 of CaBP4. Here we present NMR structures of CaBP4 in both Mg(2+)-bound and Ca(2+)-bound states and model the CaBP4 structural interaction with Cav1.4. CaBP4 contains an unstructured N-terminal region (residues 1-99) and four EF-hands in two separate lobes. The N-lobe consists of EF1 and EF2 in a closed conformation with either Mg(2+) or Ca(2+) bound at EF1. The C-lobe binds Ca(2+) at EF3 and EF4 and exhibits a Ca(2+)-induced closed-to-open transition like that of calmodulin. Exposed residues in Ca(2+)-bound CaBP4 (Phe(137), Glu(168), Leu(207), Phe(214), Met(251), Phe(264), and Leu(268)) make contacts with the IQ motif in Cav1.4, and the Cav1.4 mutant Y1595E strongly impairs binding to CaBP4. We conclude that CaBP4 forms a collapsed structure around the IQ motif in Cav1.4 that we suggest may promote channel activation by disrupting an interaction between IQ and the inhibitor of Ca(2+)-dependent inactivation domain.
钙结合蛋白 4 调节视网膜光感受器细胞中 L 型电压门控钙通道（Cav1.4）的 Ca2+依赖性活性。Mg2+结合到第一和第三 EF 手（EF1 和 EF3），而 Ca2+结合到 CaBP4 的 EF1、EF3 和 EF4。在这里，我们呈现了 CaBP4 在结合 Mg2+和 Ca2+状态下的 NMR 结构，并对 CaBP4 与 Cav1.4 的结构相互作用进行了建模。CaBP4 包含一个无结构的 N 端区域（残基 1-99）和两个独立叶中的四个 EF 手。N 叶由 EF1 和 EF2 组成，处于闭合构象，EF1 结合 Mg2+或 Ca2+。C 叶结合 EF3 和 EF4 处的 Ca2+，并表现出类似于钙调蛋白的 Ca2+-诱导的闭合-开放转变。结合 Ca2+的 CaBP4 中的暴露残基（Phe137、Glu168、Leu207、Phe214、Met251、Phe264 和 Leu268）与 Cav1.4 中的 IQ 基序接触，而 Cav1.4 突变体 Y1595E 强烈损害与 CaBP4 的结合。我们得出结论，CaBP4 在 Cav1.4 的 IQ 基序周围形成一个塌陷结构，我们认为该结构可能通过破坏 IQ 与 Ca2+依赖性失活域抑制剂之间的相互作用来促进通道激活。
