Choe Seungho, Grabe Michael
Department of Biological Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania 15260, USA.
J Chem Phys. 2009 Jun 7;130(21):215103. doi: 10.1063/1.3138906.
Voltage-gated potassium (Kv) channels control the electrical excitability of neurons and muscles. Despite this key role, how these channels open and close or gate is not fully understood. Gating is usually attributed to the bending and straightening of pore-lining helices at glycine and proline residues. In this work we focused on the role of proline in the Pro-Val-Pro (PVP) motif of the inner S6 helix in the Kv1.2 channel. We started by developing a simple hinged-rod model to fully explore the configurational space of bent helices and we related these configurations to the degree of pore opening. We then carried out fully atomistic simulations of the S6 helices and compared these simulations to the hinged-rod model. Both methods suggest that Kv1 channels are not tightly closed when the inner helices are straight, unlike what is seen in the non-PVP containing channels KcsA and KirBac. These results invite the possibility that the S6 helices may be kinked when Kv1 channels are closed. Our simulations indicate that the wild-type helix adopts multiple spatially distinct configurations, which is consistent with its role in adopting a closed state and an open state. The two most dominant configurational basins correspond to a 6 A movement of the helix tail accompanied by the PVP region undergoing a local alpha-helix to 3(10)-helix transition. We explored how single point mutations affect the propensity of the S6 helix to adopt particular configurations. Interestingly, mutating the first proline, P405 (P473 in Shaker), to alanine completely removed the bistable nature of the S6 helix possibly explaining why this mutation compromises the channel. Next, we considered four other mutations in the area known to affect channel gating and we saw similarly dramatic changes to the helix's dynamics and range of motion. Our results suggest a possible mechanism of helix pore closure and they suggest differences in the closed state of glycine-only channels, like KcsA, and PVP containing channels.
电压门控钾（Kv）通道控制着神经元和肌肉的电兴奋性。尽管发挥着这一关键作用，但这些通道如何开启和关闭或门控尚未完全明确。门控通常归因于孔道内衬螺旋在甘氨酸和脯氨酸残基处的弯曲和伸直。在这项工作中，我们聚焦于脯氨酸在Kv1.2通道内S6螺旋的脯氨酸 - 缬氨酸 - 脯氨酸（PVP）基序中的作用。我们首先构建了一个简单的铰链杆模型，以充分探索弯曲螺旋的构象空间，并将这些构象与孔道开放程度相关联。然后，我们对S6螺旋进行了全原子模拟，并将这些模拟结果与铰链杆模型进行比较。两种方法均表明，与不含PVP的通道KcsA和KirBac不同，当内部螺旋伸直时，Kv通道并非紧密关闭。这些结果提示，当Kv通道关闭时，S6螺旋可能发生扭结。我们的模拟表明，野生型螺旋会采用多种空间上不同的构象，这与其在采用关闭状态和开放状态时所起的作用相一致。两个最主要的构象盆地对应于螺旋尾部6埃的移动，同时PVP区域经历从局部α螺旋到3（10）螺旋的转变。我们探究了单点突变如何影响S6螺旋采用特定构象的倾向。有趣的是，将第一个脯氨酸P405（在Shaker中为P473）突变为丙氨酸，完全消除了S6螺旋的双稳态性质，这可能解释了为何该突变会损害通道功能。接下来，我们考虑了已知影响通道门控的该区域的其他四个突变，发现螺旋的动力学和运动范围同样发生了显著变化。我们的结果提示了一种螺旋孔道关闭的可能机制，并表明仅含甘氨酸的通道（如KcsA）与含PVP通道在关闭状态上存在差异。
