Mutations in the S4 region isolate the final voltage-dependent cooperative step in potassium channel activation.
Mutations in the S4 region isolate the final voltage-dependent cooperative step in potassium channel activation.
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DOI：
10.1085/jgp.113.3.389
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发表时间：
1999-03
影响因子：
3.8
通讯作者：
Aldrich, RW
中科院分区：
文献类型：
--
作者：
Ledwell, JL;Aldrich, RW
S4跨膜片段中的带电残基在确定电压门控离子通道对跨细胞膜电压变化的敏感性方面起关键作用。然而，亚基之间的合作相互作用也影响通道开放的电压依赖性，并且这些相互作用可以通过在S4区域中不带电荷的残基处进行取代来改变。我们已经研究了两个突变体Shaker通道的激活，它们具有不同的S4氨基酸序列，ILT（V369I，I372L和S376T）和Shaw S4（果蝇Shaw的S4取代成Shaker），但具有非常相似的离子电流特性。这两种突变都会影响协同性，使激活途径的速率限制合作过渡，并将其转移到非常正的电压，但门控和离子电流记录的分析表明，ILT和Shaw S4突变体通道具有不同的激活途径。门控电流的分析表明，ILT突变的主导作用是使最终的合作过渡到开放状态的通道速率限制的激活途径，否则类似于振动筛。与ILT激活中的最终门控转变相关联的电荷移动可以被测量为在通道打开的电压范围内的电荷移动的隔离分量，并且占ILT激活途径中移动的总电荷的13%（1.8e0）。ILT门控电荷的其余部分（87%）在负电压下移动，其中通道不打开，并证实激活途径中闭合状态之间存在Shaker样构象变化。与ILT相反，Shaw S4的激活途径似乎涉及闭合状态和开放状态之间的单个合作电荷移动步骤。我们不能检测到任何电压依赖性转换之间的关闭状态的肖S4。恢复在Shaw S4（R1、R2和K7）中缺失的碱性残基拯救了激活途径中闭合状态之间的电荷移动，但不改变激活中限速转变的电压依赖性。
Charged residues in the S4 transmembrane segment play a key role in determining the sensitivity of voltage-gated ion channels to changes in voltage across the cell membrane. However, cooperative interactions between subunits also affect the voltage dependence of channel opening, and these interactions can be altered by making substitutions at uncharged residues in the S4 region. We have studied the activation of two mutant Shaker channels that have different S4 amino acid sequences, ILT (V369I, I372L, and S376T) and Shaw S4 (the S4 of Drosophila Shaw substituted into Shaker), and yet have very similar ionic current properties. Both mutations affect cooperativity, making a cooperative transition in the activation pathway rate limiting and shifting it to very positive voltages, but analysis of gating and ionic current recordings reveals that the ILT and Shaw S4 mutant channels have different activation pathways. Analysis of gating currents suggests that the dominant effect of the ILT mutation is to make the final cooperative transition to the open state of the channel rate limiting in an activation pathway that otherwise resembles that of Shaker. The charge movement associated with the final gating transition in ILT activation can be measured as an isolated component of charge movement in the voltage range of channel opening and accounts for 13% (∼1.8 e0) of the total charge moved in the ILT activation pathway. The remainder of the ILT gating charge (87%) moves at negative voltages, where channels do not open, and confirms the presence of Shaker-like conformational changes between closed states in the activation pathway. In contrast to ILT, the activation pathway of Shaw S4 seems to involve a single cooperative charge-moving step between a closed and an open state. We cannot detect any voltage-dependent transitions between closed states for Shaw S4. Restoring basic residues that are missing in Shaw S4 (R1, R2, and K7) rescues charge movement between closed states in the activation pathway, but does not alter the voltage dependence of the rate-limiting transition in activation.
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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
影响因子：
56.9
作者：
Doyle, DA;Cabral, JM;MacKinnon, R
通讯作者：
MacKinnon, R
影响因子：
11.4
作者：
BAUMANN, A;GRUPE, A;PONGS, O
通讯作者：
PONGS, O
影响因子：
3.5
作者：
KAYANO, T;NODA, M;NUMA, S
通讯作者：
NUMA, S