A gating charge interaction required for late slow inactivation of the bacterial sodium channel NavAb.
A gating charge interaction required for late slow inactivation of the bacterial sodium channel NavAb.
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DOI：
10.1085/jgp.201311012
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发表时间：
2013-09
期刊：
影响因子：
--
通讯作者：
Catterall WA
中科院分区：
文献类型：
--
作者：
Gamal El-Din TM;Martinez GQ;Payandeh J;Scheuer T;Catterall WA
电压门控钠通道在重复去极化期间经历缓慢失活，其控制动作电位爆发的频率和持续时间并防止兴奋性毒性细胞死亡。虽然同源四聚体细菌钠通道缺乏导致真核钠通道快速失活的细胞内接头连接同源结构域III和IV，但它们保留了缓慢失活的分子机制。在这里，我们研究的功能特性和缓慢失活的细菌钠通道NavAb在昆虫细胞中表达的条件下用于结构研究。NavAb在非常负的膜电位（V1/2约为−98 mV）下激活，并且它具有在单次去极化期间出现并快速逆转的缓慢失活的早期阶段和非常缓慢逆转的缓慢失活的晚期使用依赖性阶段。在电压传感器的细胞外负簇中的S2段中Asn 49突变为Lys使激活曲线移动约75 mV至更正的电位，并消除缓慢失活的晚期。门控电荷R3与NavAb晶体结构中的Asn 49相互作用，并且该残基突变为Cys导致与突变N49 K类似的激活的电压依赖性的正向偏移和缓慢失活晚期的阻断。诱导缓慢失活的长时间去极化也会导致门控电荷运动的滞后，这导致需要非常负的膜电位才能将门控电荷返回到其静息状态。出乎意料的是，突变N49 K不改变门控电荷运动的滞后，即使它阻止了缓慢失活的晚期。我们的研究结果揭示了S4中的R3和S2中的Asn 49之间的重要分子相互作用，这对于NavAb的电压依赖性激活和晚期缓慢失活至关重要，并且它们引入了NavAb突变体，该突变体能够与结构分析并行进行详细的功能研究。
Voltage-gated sodium channels undergo slow inactivation during repetitive depolarizations, which controls the frequency and duration of bursts of action potentials and prevents excitotoxic cell death. Although homotetrameric bacterial sodium channels lack the intracellular linker-connecting homologous domains III and IV that causes fast inactivation of eukaryotic sodium channels, they retain the molecular mechanism for slow inactivation. Here, we examine the functional properties and slow inactivation of the bacterial sodium channel NavAb expressed in insect cells under conditions used for structural studies. NavAb activates at very negative membrane potentials (V1/2 of approximately −98 mV), and it has both an early phase of slow inactivation that arises during single depolarizations and reverses rapidly, and a late use-dependent phase of slow inactivation that reverses very slowly. Mutation of Asn49 to Lys in the S2 segment in the extracellular negative cluster of the voltage sensor shifts the activation curve ∼75 mV to more positive potentials and abolishes the late phase of slow inactivation. The gating charge R3 interacts with Asn49 in the crystal structure of NavAb, and mutation of this residue to Cys causes a similar positive shift in the voltage dependence of activation and block of the late phase of slow inactivation as mutation N49K. Prolonged depolarizations that induce slow inactivation also cause hysteresis of gating charge movement, which results in a requirement for very negative membrane potentials to return gating charges to their resting state. Unexpectedly, the mutation N49K does not alter hysteresis of gating charge movement, even though it prevents the late phase of slow inactivation. Our results reveal an important molecular interaction between R3 in S4 and Asn49 in S2 that is crucial for voltage-dependent activation and for late slow inactivation of NavAb, and they introduce a NavAb mutant that enables detailed functional studies in parallel with structural analysis.
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影响因子：
64.8
作者：
Payandeh, Jian;El-Din, Tamer M. Gamal;Scheuer, Todd;Zheng, Ning;Catterall, William A.
通讯作者：
Catterall, William A.
DOI：
10.1073/pnas.0806486105
发表时间：
2008-09-30
影响因子：
11.1
作者：
DeCaen, Paul G.;Yarov-Yarovoy, Vladimir;Catterall, William A.
通讯作者：
Catterall, William A.
DOI：
10.1085/jgp.201010573
发表时间：
2011-05
期刊：
The Journal of general physiology
影响因子：
--
作者：
Haddad GA;Blunck R
通讯作者：
Blunck R
影响因子：
3.4
作者：
Pavlov, E;Bladen, C;French, RJ
通讯作者：
French, RJ
影响因子：
64.8
作者：
通讯作者：
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