Restoration of fast inactivation in an inactivation-defective human heart sodium channel by the cysteine modifying reagent Benzyl-MTS: Analysis of IFM-ICM mutation
Restoration of fast inactivation in an inactivation-defective human heart sodium channel by the cysteine modifying reagent Benzyl-MTS: Analysis of IFM-ICM mutation
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DOI：
10.1006/bbrc.1997.6510
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发表时间：
1997-04-28
影响因子：
3.1
通讯作者：
Kallen, RG
中科院分区：
文献类型：
--
作者：
Chahine, M;Deschenes, I;Kallen, RG
有人认为，连接电压门控钠通道结构域III和IV的区域形成了失活门。结合定点突变、半胱氨酸共价修饰和电生理记录技术来确定Phe(1486)的作用，Phe(1486)是位于Na+通道的III-IV连接子上的一个保守的苯丙氨酸残基，它是疏水氨基酸簇(IFM)的一部分，被认为在电压门控钠通道的快速失活中起重要作用。在改变的人类心脏1 Na+通道(HHI/F14SGC)中，Phe(1486)被半胱氨酸取代的tsA201细胞中的表达与残余电流的出现、失活时间常数的电压依赖性的丧失、稳态失活向更去极化的电压的转变以及比野生型Hill更快的失活恢复有关。突变F1486C的细胞质表面暴露于甲硫磺酸试剂MTSEA、MTSET和MTSES，进一步破坏了宏观失活，但暴露于MTSBN完全恢复了快速失活和快速失活的电压依赖性。这些发现支持电压门控钠通道III-IV连接子的IFM基序是失活粒子的必要成分，并且Phe(1486)的苯基可能在失活门关闭中起关键作用。(C)1997年学术出版社。
It has been suggested that the region linking domain III and IV of voltage-gated sodium channels forms the inactivation gate. A combination of site-directed mutagenesis, cysteine covalent modification, and electrophysiological recording techniques was used to identify the role of the Phe(1486), a conserved phenylalanine residue located in the III-IV linker of Na+ channels, This Phe(1486) is part of a hydrophobic amino acid cluster (IFM) that was proposed to play an essential role in the fast inactivation of voltage-gated sodium channels. Expression in tsA201 cells of an altered human heart 1 Na+ channel (hHI/F14SGC) in which Phe(1486) was replaced by a cysteine is associated with the appearance of a residual current, a loss of voltage-dependence of the time constants of inactivation, a shift of the steady-state inactivation to more depolarized voltages, and a recovery from inactivation that is faster than the wild-type hill. Exposure of the cytoplasmic surface of mutant F1486C to the methanthiosulfonate reagents, MTSEA, MTSET, and MTSES, further disrupted macroscopic inactivation, but exposure to MTSBN completely restores fast inactivation and the voltage-dependence of fast inactivation. These findings support the formulation that the IFM motif of the III-IV-linker of voltage-gated sodium channels serves as an essential component of the inactivation particle and that the phenyl group of Phe(1486) may play a crucial role in inactivation gate closure. (C) 1997 Academic Press.