Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, Illinois; Department of Physiology and Biophysics, Weill Cornell Medical College, New York, New York.
Department of Physiology and Biophysics, Weill Cornell Medical College, New York, New York.
Biophys J. 2012 Aug 8;103(3):434-443. doi: 10.1016/j.bpj.2012.06.023.
Inward rectifier potassium (Kir) channels act as cellular diodes, allowing unrestricted flow of potassium (K(+)) into the cell while preventing currents of large magnitude in the outward direction. The rectification mechanism by which this occurs involves a coupling between K(+) and intracellular blockers-magnesium (Mg(2+)) or polyamines-that simultaneously occupy the permeation pathway. In addition to the transmembrane pore, Kirs possess a large cytoplasmic domain (CD) that provides a favorable electronegative environment for cations. Electrophysiological experiments have shown that the CD is a key regulator of both conductance and rectification. In this study, we calculate and compare averaged equilibrium probability densities of K(+) and Cl(-) in open-pore models of the CDs of a weak (Kir1.1-ROMK) and a strong (Kir2.1-IRK) rectifier through explicit-solvent molecular-dynamics simulations in ~1 M KCl. The CD of both channels concentrates K(+) ions greater than threefold inside the cytoplasmic pore while IRK shows an additional K(+) accumulation region near the cytoplasmic entrance. Simulations carried out with Mg(2+) or spermine (SPM(4+)) show that these ions interact with pore-lining residues, shielding the surface charge and reducing K(+) in both channels. The results also show that SPM(4+) behaves differently inside these two channels. Although SPM(4+) remains inside the CD of ROMK, it diffuses around the entire volume of the pore. In contrast, this polyatomic cation finds long-lived conformational states inside the IRK pore, interacting with residues E224, D259, and E299. The strong rectifier CD is also capable of sequestering an additional SPM(4+) at the cytoplasmic entrance near a cluster of negative residues D249, D274, E275, and D276. Although understanding the actual mechanism of rectification blockade will require high-resolution structural information of the blocked state, these simulations provide insight into how sequence variation in the CD can affect the multi-ion distributions that underlie the mechanisms of conduction, rectification affinity, and kinetics.
内向整流钾 (Kir) 通道作为细胞二极管，允许钾 (K(+)) 无限制地流入细胞，同时防止向外的大电流。发生这种整流的机制涉及 K(+) 和细胞内阻断剂-镁 (Mg(2+)) 或聚胺-的同时占据渗透途径的耦合。除了跨膜孔外，Kir 还具有大的细胞质结构域 (CD)，为阳离子提供有利的电负性环境。电生理实验表明，CD 是电导和整流的关键调节剂。在这项研究中，我们通过在 ~1 M KCl 中进行显式溶剂分子动力学模拟，计算并比较了弱 (Kir1.1-ROMK) 和强 (Kir2.1-IRK) 整流器 CD 的开放孔模型中 K(+) 和 Cl(-) 的平均平衡概率密度。两个通道的 CD 在细胞质孔内将 K(+) 离子浓缩超过三倍，而 IRK 在细胞质入口附近显示出额外的 K(+) 积累区域。用镁 (Mg(2+)) 或 spermine (SPM(4+)) 进行的模拟表明，这些离子与孔衬里残基相互作用，屏蔽表面电荷并减少两个通道中的 K(+)。结果还表明，SPM(4+) 在这两个通道中的行为不同。虽然 SPM(4+) 留在 ROMK 的 CD 内，但它在整个孔体积内扩散。相比之下，这种多原子阳离子在 IRK 孔内找到长寿命的构象状态，与残基 E224、D259 和 E299 相互作用。强整流器 CD 还能够在细胞质入口附近的一组负电荷残基 D249、D274、E275 和 D276 处隔离额外的 SPM(4+)。尽管了解整流阻断的实际机制将需要阻塞状态的高分辨率结构信息，但这些模拟提供了对 CD 中的序列变异如何影响传导、整流亲和力和动力学机制下的多离子分布的深入了解。
