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Identification of the activation gate of HERG K^+ channel

HERG K^通道激活门的识别

基本信息

批准号：
14370028
负责人：
ISHII Kuniaki
金额：
$ 8.06万
依托单位：
Yamagata University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2002
资助国家：
日本
起止时间：
2002 至 2004
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-14370028/
关键词：
HERG voltage sensor activation gate outward currents inward currents ion selectivity charged amino acids 第6膜貫通領域

项目摘要

1. Wild type HERG K^+ channel opens upon depolarization and does not open upon hyperpolarization. We have found that a point mutation at I655 on the S6 makes the HERG channel open upon hyperpolarization. Further mutagenesis analyses showed that substitution of I655 with charged or polar amino acid residues resulted in the similar change (The resultant mutants opened upon hyperpolarization). On the other hand, HERG K^+ channel seemed to require hydrophobic residues at 655 to open upon depolarization like the wild type channel.2. Similar but not identical results were obtained with point mutations of G657 on the S6. When charged residues (except glutamatic acid) or polar residues were present at 657, the mutant channels opened upon hyperpolarization. In the case of the residue at 657, HERG K^+ channel seemed to require a small residue such as alanine or serine at 657 to open upon depolarization.3. Selectivity for K^+ ion was lost in the I655 and G657 mutant channels that open upon hyperpolarization. Although the mechanism is not known, opening of the mutant channels upon hyperpolarization seemed to involve the movement of the ion selectivity filter.4. When the three charged residues on the S4-S5 linker of HERG WT channel were neutralized, upon hyperpolarization, obvious inward currents flowing through the mutant channel were observed, in addition to outward currents on depolarization. However, the neutralization of the charged residues did not cause any change in the I655 and G657 mutants that open upon hyperpolarization.5. It has been reported that the inner helices bend at the glycine hinge, when K^+ channel opens. Substitution of the corresponding glycine in HERG with alanine suggested that the glycine hinge is involved in the opening of the WT HERG channel, but not in the I655 and G657 mutant channels.

1.野生型HERG K^+通道在去极化时开放，在超极化时不开放。我们已经发现，S6上I655的点突变使HERG通道在超极化时打开。进一步的突变分析表明，用带电荷的氨基酸残基或极性氨基酸残基替换I655会导致类似的变化(所产生的突变体在超极化时打开)。另一方面，HERG K^+通道似乎需要655位疏水残基才能像野生型通道那样在去极化时开放。S6上G657的点突变也得到了相似但不相同的结果。当在657处存在带电残基(谷氨酸除外)或极性残基时，突变通道在超极化时打开。在657位残基的情况下，Herg K^+通道似乎需要657位的小残基，如丙氨酸或丝氨酸，才能在去极化时打开。在超极化时开放的I655和G657突变通道中，K~(++)的选择性丧失。虽然机制尚不清楚，但超极化时突变通道的打开似乎涉及到离子选择性过滤器的移动。当HERG WT通道S4-S5连接子上的三个带电残基被中和时，在超极化时，除了去极化的外向电流外，还观察到明显的内向电流流过突变通道。然而，带电残基的中和并没有引起I655和G657突变体在超极化时打开的任何变化。据报道，当K^+通道开放时，内螺旋在甘氨酸铰链处弯曲。用丙氨酸取代HERG中相应的甘氨酸，表明甘氨酸铰链参与了WT HERG通道的开放，但不参与I655和G657突变通道的开放。