Direct Regulation of Prokaryotic Kir Channel by Cholesterol
Direct Regulation of Prokaryotic Kir Channel by Cholesterol
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DOI：
10.1074/jbc.m109.011221
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发表时间：
2009-10-30
影响因子：
4.8
通讯作者：
Levitan, Irena
中科院分区：
文献类型：
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作者：
Singh, Dev K.;Rosenhouse-Dantsker, Avia;Levitan, Irena
我们早期的研究表明，细胞胆固醇的升高强烈抑制了内向整流器Kir 2亚家族的通道活性。本研究的目的是确定胆固醇是否直接抑制Kir通道。为了实现这一目标，将纯化的原核Kir（KirBac1.1）通道掺入到限定脂质组成的脂质体中，并通过Rb-86（+）摄取测定通道活性。我们的研究结果表明，Rb-86（+）通过KirBac1.1通量强烈抑制胆固醇。将5%（质量胆固醇/磷脂）胆固醇掺入脂质体中抑制Rb-86（+）通量> 50%，并且活性在12- 15%时被完全抑制。然而，表胆固醇，一种具有相似物理性质的胆固醇的立体异构体，对KirBac介导的Rb-86（+）摄取的影响明显小于胆固醇。此外，多种甾醇的分析表明，胆固醇诱导的抑制KirBac1.1通道介导的特定的相互作用，而不是通过改变脂质双层的物理性质。与KirBac1.1活性的抑制相反，胆固醇对重建的KscA通道的活性没有影响（高达250 μ g/mg磷脂）。总之，这些观察结果表明，胆固醇抑制Kir通道在一个纯的蛋白质-脂质环境，并表明，相互作用是直接和具体的。
Our earlier studies have shown that channel activity of Kir2 subfamily of inward rectifiers is strongly suppressed by the elevation of cellular cholesterol. The goal of this study is to determine whether cholesterol suppresses Kir channels directly. To achieve this goal, purified prokaryotic Kir (KirBac1.1) channels were incorporated into liposomes of defined lipid composition, and channel activity was assayed by Rb-86(+) uptake. Our results show that Rb-86(+) flux through KirBac1.1 is strongly inhibited by cholesterol. Incorporation of 5% ( mass cholesterol/phospholipid) cholesterol into the liposome suppresses Rb-86(+) flux by >50%, and activity is completely inhibited at 12-15%. However, epicholesterol, a stereoisomer of cholesterol with similar physical properties, has significantly less effect on KirBac-mediated Rb-86(+) uptake than cholesterol. Furthermore, analysis of multiple sterols suggests that cholesterol-induced inhibition of KirBac1.1 channels is mediated by specific interactions rather than by changes in the physical properties of the lipid bilayer. In contrast to the inhibition of KirBac1.1 activity, cholesterol had no effect on the activity of reconstituted KscA channels (at up to 250 mu g/mg of phospholipid). Taken together, these observations demonstrate that cholesterol suppresses Kir channels in a pure protein-lipid environment and suggest that the interaction is direct and specific.