KCNE peptides differently affect voltage sensor equilibrium and equilibration rates in KCNQ1 K+ channels.
KCNE peptides differently affect voltage sensor equilibrium and equilibration rates in KCNQ1 K+ channels.
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DOI：
10.1085/jgp.200709816
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发表时间：
2008-01
期刊：
影响因子：
--
通讯作者：
Kobertz WR
中科院分区：
文献类型：
--
作者：
Rocheleau JM;Kobertz WR
KCNQ 1电压门控K+通道与KCNE I型跨膜肽家族组装，以提供具有不同通道门控特性的膜包埋复合物。KCNQ 1/KCNE 1复合物产生非常缓慢激活的心脏IKs电流，而与KCNE 3组装产生参与上皮细胞中K+再循环的组成型传导复合物。为了确定这两个KCNE肽是否影响KCNQ 1通道中的电压传感，我们使用半胱氨酸可及性实验监测了KCNQ 1/KCNE复合物中S4电压传感器的位置。一组KCNQ 1 S4半胱氨酸突变体在非洲爪蟾卵母细胞中表达，用膜不渗透半胱氨酸特异性试剂2-（三甲基铵）乙基甲硫基磺酸盐（MTSET）处理，并确定每个突变体的电压依赖性可及性。在这些S4半胱氨酸突变体中，只有当KCNQ 1去极化时，MTSET才能修饰三个突变体（R228 C，G229 C，I230 C）。然后，我们采用这些状态依赖的残基，以确定如何组装KCNE 1和KCNE 3影响KCNQ 1电压传感器的平衡和平衡速率。在KCNE 1存在下，大多数半胱氨酸突变体的MTSET修饰速率慢10倍，如最近报道的，表明KCNE 1使KCNQ 1电压传感器的动力学减慢（Nakajo，K.，和Y.久保130：269-281）。由于MTS修饰率反映了试剂可及性，化学反应性和蛋白质构象变化的混合物，我们改变了去极化脉冲持续时间，以确定KCNE 1是否减慢电压传感器的平衡速率。使用状态依赖的半胱氨酸突变体，我们确定MTSET修饰率基本上是独立的去极化脉冲持续时间。这些结果表明，在去极化时，电压传感器在KCNE 1的存在下快速达到平衡，并且通道复合物的缓慢门控不是由于缓慢移动的电压传感器。与此相反，KCNQ 1/KCNE 3复合物的S4中的所有半胱氨酸取代都可以自由地接近MTSET，而不受电压的影响，这与KCNE 3使电压传感器平衡移动以有利于超极化电位下的活性状态是一致的。总之，这些结果表明KCNE肽不同地调节KCNQ 1 K+通道中的电压传感器。
KCNQ1 voltage-gated K+ channels assemble with the family of KCNE type I transmembrane peptides to afford membrane-embedded complexes with diverse channel gating properties. KCNQ1/KCNE1 complexes generate the very slowly activating cardiac IKs current, whereas assembly with KCNE3 produces a constitutively conducting complex involved in K+ recycling in epithelia. To determine whether these two KCNE peptides influence voltage sensing in KCNQ1 channels, we monitored the position of the S4 voltage sensor in KCNQ1/KCNE complexes using cysteine accessibility experiments. A panel of KCNQ1 S4 cysteine mutants was expressed in Xenopus oocytes, treated with the membrane-impermeant cysteine-specific reagent 2-(trimethylammonium) ethyl methanethiosulfonate (MTSET), and the voltage-dependent accessibility of each mutant was determined. Of these S4 cysteine mutants, three (R228C, G229C, I230C) were modified by MTSET only when KCNQ1 was depolarized. We then employed these state-dependent residues to determine how assembly with KCNE1 and KCNE3 affects KCNQ1 voltage sensor equilibrium and equilibration rates. In the presence of KCNE1, MTSET modification rates for the majority of the cysteine mutants were ∼10-fold slower, as was recently reported to indicate that the kinetics of the KCNQ1 voltage sensor are slowed by KCNE1 (Nakajo, K., and Y. Kubo. 2007 J. Gen. Physiol. 130:269–281). Since MTS modification rates reflect an amalgam of reagent accessibility, chemical reactivity, and protein conformational changes, we varied the depolarization pulse duration to determine whether KCNE1 slows the equilibration rate of the voltage sensors. Using the state-dependent cysteine mutants, we determined that MTSET modification rates were essentially independent of depolarization pulse duration. These results demonstrate that upon depolarization the voltage sensors reach equilibrium quickly in the presence of KCNE1 and the slow gating of the channel complex is not due to slowly moving voltage sensors. In contrast, all cysteine substitutions in the S4 of KCNQ1/KCNE3 complexes were freely accessible to MTSET independent of voltage, which is consistent with KCNE3 shifting the voltage sensor equilibrium to favor the active state at hyperpolarizing potentials. In total, these results suggest that KCNE peptides differently modulate the voltage sensor in KCNQ1 K+ channels.
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影响因子：
16.2
作者：
Aggarwal, SK;MacKinnon, R
通讯作者：
MacKinnon, R
影响因子：
16.2
作者：
Larsson, HP;Baker, OS;Isacoff, EY
通讯作者：
Isacoff, EY
振动弹器K+通道中电压传感器运动的限制。
影响因子：
3.8
作者：
Darman, Rachel B;Ivy, Allison A;Ketty, Vina;Blaustein, Robert O
通讯作者：
Blaustein, Robert O
影响因子：
64.8
作者：
Männikkö, R;Elinder, F;Larsson, HP
通讯作者：
Larsson, HP
影响因子：
5.5
作者：
Panaghie, G;Tai, KK;Abbott, GW
通讯作者：
Abbott, GW