Two arginines in the cytoplasmic C-terminal domain are essential for voltage-dependent regulation of A-type K+ current in the Kv4 channel subfamily
Two arginines in the cytoplasmic C-terminal domain are essential for voltage-dependent regulation of A-type K+ current in the Kv4 channel subfamily
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DOI：
10.1074/jbc.m302034200
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发表时间：
2004-02-13
影响因子：
4.8
通讯作者：
Imaizumi, Y
中科院分区：
文献类型：
--
作者：
Hatano, N;Ohya, S;Imaizumi, Y
Kv4.3的C端结构域对A型K+电流（I-A）的电压依赖性门控的贡献通过（i）在该区域进行突变，（ii）在HEK 293细胞中的异源表达和（iii）详细的电压钳分析来检查。大鼠Kv4.3M的C末端的进行性缺失（从N末端到氨基酸429）没有显著改变IA的失活时间过程，但将稳态失活的电压依赖性向负方向移动至最大值-17 mV。进一步的缺失（氨基酸420）转移这个参数在积极的方向，这表明在IA的电压依赖性调节域429-420的关键作用。在该结构域中有四个带正电荷的氨基酸：Lys（423）、Lys（424）、Arg（426）和Arg（429）。用丙氨酸（R2 A）替换两种精氨酸分别导致失活和活化的-23和-13 mV偏移。用丙氨酸额外替换两个赖氨酸没有导致进一步的移位。R426 A或R429 A的单次置换分别诱导-15和-10 mV的失活偏移。R2 A没有显着改变失活率，但显着改变失活恢复的电压依赖性。这两个精氨酸在Kv 4亚家族中是保守的，并且在Kv4.2中Arg（429）和Arg（432）的丙氨酸替换得到基本上相同的结果。R2 A的这些作用不受K+通道β亚基KChIPs共表达的调节。总之，Kv 4亚家族α-亚基的胞质C-末端结构域中的两个丝氨酸强烈调节通道激活、失活和恢复的电压依赖性。
Contributions of the C-terminal domain of Kv4.3 to the voltage-dependent gating of A-type K+ current (I-A) were examined by (i) making mutations in this region, (ii) heterologous expression in HEK293 cells, and (iii) detailed voltage clamp analyses. Progressive deletions of the C terminus of rat Kv4.3M ( to amino acid 429 from the N terminus) did not markedly change the inactivation time course of IA but shifted the voltage dependence of steady state inactivation in the negative direction to a maximum of -17 mV. Further deletions ( to amino acid 420) shifted this parameter in the positive direction, suggesting a critical role for the domain 429-420 in the voltage-dependent regulation of IA. There are four positively charged amino acids in this domain: Lys(423), Lys(424), Arg(426), and Arg(429). The replacement of the two arginines with alanines (R2A) resulted in -23 and -13 mV shifts of inactivation and activation, respectively. Additional replacement of the two lysines with alanines did not result in further shifts. Single replacements of R426A or R429A induced -15 and -10 mV shifts of inactivation, respectively. R2A did not significantly change the inactivation rate but did markedly change the voltage dependence of recovery from inactivation. These two arginines are conserved in Kv4 subfamily, and alanine replacement of Arg(429) and Arg(432) in Kv4.2 gave essentially the same results. These effects of R2A were not modulated by co-expression of the K+ channel beta subunit, KChIPs. In conclusion, the two arginines in the cytosolic C-terminal domain of alpha-subunits of Kv4 subfamily strongly regulate the voltage dependence of channel activation, inactivation, and recovery.