Ellinor P T, Yang J, Sather W A, Zhang J F, Tsien R W
Department of Molecular and Cellular Physiology, Beckman Center, Stanford University Medical Center, California 94305, USA.
Neuron. 1995 Nov;15(5):1121-32. doi: 10.1016/0896-6273(95)90100-0.
Ca2+ channels display remarkable selectivity and permeability, traditionally attributed to multiple, discrete Ca2+ binding sites lining the pore. Each of the four pore-forming segments of Ca2+ channel alpha 1 subunits contains a glutamate residue that contributes to high-affinity Ca2+ interactions. Replacement of all four P-region glutamates with glutamine or alanine abolished micromolar Ca2+ block of monovalent current without revealing any additional independent high-affinity Ca2+ binding site. Pairwise replacements of the four glutamates excluded the hypothesis that they form two independent high-affinity sites. Systematic alterations of side-chain length, charge, and polarity by glutamate replacement with aspartate, glutamine, or alanine weakened the Ca2+ interaction, with considerable asymmetry from one repeat to another. The P-region glutamate in repeat I was unusual in its sensitivity to aspartate replacement but not glutamine substitution. While all four glutamates cooperate in supporting high-affinity interactions with single Ca2+ ions, they also influence the interaction between multiple divalent cations.
钙离子通道表现出显著的选择性和通透性，传统上认为这归因于孔道内衬的多个离散钙离子结合位点。钙离子通道α1亚基的四个孔形成片段中的每一个都含有一个谷氨酸残基，这有助于形成高亲和力的钙离子相互作用。用谷氨酰胺或丙氨酸取代所有四个P区谷氨酸消除了微摩尔浓度钙离子对单价电流的阻断，且未揭示任何额外的独立高亲和力钙离子结合位点。四个谷氨酸的成对替换排除了它们形成两个独立高亲和力位点的假说。通过用天冬氨酸、谷氨酰胺或丙氨酸取代谷氨酸来系统改变侧链长度、电荷和极性，削弱了钙离子相互作用，不同重复片段之间存在相当大的不对称性。重复片段I中的P区谷氨酸对天冬氨酸取代的敏感性不同寻常，但对谷氨酰胺取代不敏感。虽然所有四个谷氨酸协同支持与单个钙离子的高亲和力相互作用，但它们也影响多个二价阳离子之间的相互作用。
