NMR-derived dynamic aspects of N-type inactivation of a Kv channel suggest a transient interaction with the T1 domain
NMR-derived dynamic aspects of N-type inactivation of a Kv channel suggest a transient interaction with the T1 domain
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DOI：
10.1021/bi0516430
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发表时间：
2006-02-14
期刊：
影响因子：
2.9
通讯作者：
Choe, S
中科院分区：
文献类型：
--
作者：
Baker, KA;Hilty, C;Choe, S
一些真核细胞电压门控K+（Kv）通道含有N-末端失活肽，其介导在膜去极化期间限制通道功能的快速失活过程，从而形成动作电位。我们获得了序列特异性核磁共振（NMR）分配的多肽骨架的四聚体N-末端片段（氨基酸1-181）的Aesthesia Kv1.1通道。另外的NMR测量显示，四聚结构域1（T1）在溶液中具有与先前通过晶体学确定的相同的球状结构，并且IP（残基1-20）和接头（残基21-65）处于柔性无序的、主要延伸的构象。T1结构域和柔性尾（残基1 - 65）之间的潜在接触位点已经基于单个T1结构域氨基酸的化学位移变化被鉴定，其映射到邻近亚基之间的界面附近的T1表面。顺磁微扰实验进一步表明，在合奏的解决方案构象，有至少一个小的人口的物种与IP本地化的T1四聚体的建议相互作用的残基附近。电生理学测量表明，我们测试的这个口袋中的所有三个突变减缓了失活速率并加快了恢复，正如从预失活位点模型预测的那样。这些结果表明，特定的，短暂的短暂的T1结构域和IP或连接片段之间的相互作用可能发挥作用，在定义快通道失活的调节动力学。
Some eukaryotic voltage-gated K+ (Kv) channels contain an N-terminal inactivation peptide JP), which mediates a fast inactivation process that limits channel function during membrane depolarization and thus shapes the action potential. We obtained sequence-specific nuclear magnetic resonance (NMR) assignments for the polypeptide backbone of a tetrameric N-terminal fragment (amino acids 1-181) of the Aplysia Kv1.1 channel. Additional NMR measurements show that the tetramerization domain 1 (T1) has the same globular structure in solution as previously determined by crystallography and that the IP (residues 1-20) and the linker (residues 21-65) are in a flexibly disordered, predominantly extended conformation. A potential contact site between the T1 domain and the flexible tail (residues 1 -65) has been identified on the basis of chemical-shift changes of individual T1 domain amino acids, which map to the T1 surface near the interface between adjacent subunits. Paramagnetic perturbation experiments further indicate that, in the ensemble of solution conformers, there is at least a small population of species with the IP localized in close proximity to the proposed interacting residues of the T1 tetramer. Electrophysiological measurements show that all three mutations in this pocket that we tested slow the rate of inactivation and speed up recovery, as predicted from the preinactivation site model. These results suggest that specific, short-lived transient interactions between the T1 domain and the IP or the linker segment may play a role in defining the regulatory kinetics of fast channel inactivation.