当前位置:
X-MOL 学术
›
Biophys. J.
›
论文详情
Our official English website, www.x-mol.net, welcomes your feedback! (Note: you will need to create a separate account there.)
GsMTx4: Mechanism of Inhibiting Mechanosensitive Ion Channels
Biophysical Journal ( IF 3.2 ) Pub Date : 2017-01-01 , DOI: 10.1016/j.bpj.2016.11.013 Radhakrishnan Gnanasambandam 1 , Chiranjib Ghatak 2 , Anthony Yasmann 3 , Kazuhisa Nishizawa 4 , Frederick Sachs 1 , Alexey S Ladokhin 2 , Sergei I Sukharev 3 , Thomas M Suchyna 1
Biophysical Journal ( IF 3.2 ) Pub Date : 2017-01-01 , DOI: 10.1016/j.bpj.2016.11.013 Radhakrishnan Gnanasambandam 1 , Chiranjib Ghatak 2 , Anthony Yasmann 3 , Kazuhisa Nishizawa 4 , Frederick Sachs 1 , Alexey S Ladokhin 2 , Sergei I Sukharev 3 , Thomas M Suchyna 1
Affiliation
GsMTx4 is a spider venom peptide that inhibits cationic mechanosensitive channels (MSCs). It has six lysine residues that have been proposed to affect membrane binding. We synthesized six analogs with single lysine-to-glutamate substitutions and tested them against Piezo1 channels in outside-out patches and independently measured lipid binding. Four analogs had ∼20% lower efficacy than the wild-type (WT) peptide. The equilibrium constants calculated from the rates of inhibition and washout did not correlate with the changes in inhibition. The lipid association strength of the WT GsMTx4 and the analogs was determined by tryptophan autofluorescence quenching and isothermal calorimetry with membrane vesicles and showed no significant differences in binding energy. Tryptophan fluorescence-quenching assays showed that both WT and analog peptides bound superficially near the lipid-water interface, although analogs penetrated deeper. Peptide-lipid association, as a function of lipid surface pressure, was investigated in Langmuir monolayers. The peptides occupied a large fraction of the expanded monolayer area, but that fraction was reduced by peptide expulsion as the pressure approached the monolayer-bilayer equivalence pressure. Analogs with compromised efficacy had pressure-area isotherms with steeper slopes in this region, suggesting tighter peptide association. The pressure-dependent redistribution of peptide between "deep" and "shallow" binding modes was supported by molecular dynamics (MD) simulations of the peptide-monolayer system under different area constraints. These data suggest a model placing GsMTx4 at the membrane surface, where it is stabilized by the lysines, and occupying a small fraction of the surface area in unstressed membranes. When applied tension reduces lateral pressure in the lipids, the peptides penetrate deeper acting as "area reservoirs" leading to partial relaxation of the outer monolayer, thereby reducing the effective magnitude of stimulus acting on the MSC gate.
中文翻译:
GsMTx4:抑制机械敏感离子通道的机制
GsMTx4 是一种蜘蛛毒肽,可抑制阳离子机械敏感通道 (MSC)。它有六个赖氨酸残基,已被认为会影响膜结合。我们合成了六种具有单一赖氨酸至谷氨酸取代的类似物,并针对外侧斑块中的 Piezo1 通道对它们进行了测试,并独立测量了脂质结合。四种类似物的功效比野生型 (WT) 肽低约 20%。根据抑制率和洗脱率计算的平衡常数与抑制变化不相关。 WT GsMTx4 和类似物的脂质结合强度通过色氨酸自发荧光猝灭和膜囊泡等温量热法测定,并且显示结合能没有显着差异。色氨酸荧光猝灭测定表明,WT 和类似肽均在脂质-水界面附近表面结合,尽管类似物渗透得更深。在朗缪尔单层中研究了肽-脂质结合作为脂质表面压力的函数。肽占据了扩展的单层区域的大部分,但是当压力接近单层-双层等效压力时,该部分因肽排出而减少。功效受损的类似物在该区域的压力区域等温线具有更陡的斜率,表明肽关联更紧密。肽单层系统在不同面积限制下的分子动力学(MD)模拟支持了肽在“深”和“浅”结合模式之间的压力依赖性重新分布。这些数据表明一个模型将 GsMTx4 放置在膜表面,在那里它被赖氨酸稳定,并占据无应力膜表面积的一小部分。 当施加的张力降低脂质中的侧向压力时,肽渗透得更深,充当“区域储库”,导致外部单层部分松弛,从而降低作用于 MSC 门的有效刺激强度。
更新日期:2017-01-01
中文翻译:
![](https://scdn.x-mol.com/jcss/images/paperTranslation.png)
GsMTx4:抑制机械敏感离子通道的机制
GsMTx4 是一种蜘蛛毒肽,可抑制阳离子机械敏感通道 (MSC)。它有六个赖氨酸残基,已被认为会影响膜结合。我们合成了六种具有单一赖氨酸至谷氨酸取代的类似物,并针对外侧斑块中的 Piezo1 通道对它们进行了测试,并独立测量了脂质结合。四种类似物的功效比野生型 (WT) 肽低约 20%。根据抑制率和洗脱率计算的平衡常数与抑制变化不相关。 WT GsMTx4 和类似物的脂质结合强度通过色氨酸自发荧光猝灭和膜囊泡等温量热法测定,并且显示结合能没有显着差异。色氨酸荧光猝灭测定表明,WT 和类似肽均在脂质-水界面附近表面结合,尽管类似物渗透得更深。在朗缪尔单层中研究了肽-脂质结合作为脂质表面压力的函数。肽占据了扩展的单层区域的大部分,但是当压力接近单层-双层等效压力时,该部分因肽排出而减少。功效受损的类似物在该区域的压力区域等温线具有更陡的斜率,表明肽关联更紧密。肽单层系统在不同面积限制下的分子动力学(MD)模拟支持了肽在“深”和“浅”结合模式之间的压力依赖性重新分布。这些数据表明一个模型将 GsMTx4 放置在膜表面,在那里它被赖氨酸稳定,并占据无应力膜表面积的一小部分。 当施加的张力降低脂质中的侧向压力时,肽渗透得更深,充当“区域储库”,导致外部单层部分松弛,从而降低作用于 MSC 门的有效刺激强度。