Alternative membrane protein conformations in alcohols
Alternative membrane protein conformations in alcohols
复制标题
DOI：
10.1021/bi700091r
复制
发表时间：
2007-04-10
期刊：
影响因子：
2.9
通讯作者：
Nesgaard, L. W.
中科院分区：
文献类型：
--
作者：
Otzen, D. E.;Sehgal, P.;Nesgaard, L. W.
醇类调节脂质双分子层中膜蛋白的寡聚化。这可以通过以与醇疏水性相关的方式重新分配侧膜压力间接发生。在这里，我们研究了不同的酒精-水混合物对膜蛋白稳定性和溶解度的直接影响，使用两种洗涤剂可溶解的α -螺旋膜蛋白DsbB和NhaA。这两种蛋白质在中等浓度的醇下广泛沉淀，形成广泛（40-60%）的β -片结构和对纤维特异性染料硫黄素T的亲和力，尽管原子力显微镜图像显示层状和球形沉积物，可能是纤维过程的早期阶段，被强疏水接触所捕获。在较高的酒精浓度下，DsbB和NhaA都被分解并形成非天然结构，与天然状态相比，DsbB的螺旋度增加或NhaA的螺旋度减少。替代构象状态不能在酒精稀释后恢复到功能的天然状态。在低醇浓度下，DsbB的沉淀效率和失稳程度与醇疏水性表现出相同的相关性。因此，除了对膜的作用外，醇还通过溶剂化膜蛋白的疏水区域直接干扰膜蛋白。在中等浓度下，这种扰动暴露了疏水段，但没有提供足够的溶剂化来避免分子间的结合。再溶解需要将相对介电常数降低到65以下，并结合单个醇的特定性质。我们得出的结论是，醇类为膜蛋白提供了多种构象的途径，但并不先验地适合于需要单体蛋白可逆变性的溶液研究。
Alcohols modulate the oligomerization of membrane proteins in lipid bilayers. This can occur indirectly by redistributing lateral membrane pressure in a manner which correlates with alcohol hydrophobicity. Here we investigate the direct impact of different alcohol-water mixtures on membrane protein stability and solubility, using the two detergent-solubilized alpha-helical membrane proteins DsbB and NhaA. Both proteins precipitate extensively at intermediate concentrations of alcohols, forming states with extensive (40-60%) beta-sheet structure and affinity for the fibril-specific dye thioflavin T, although atomic force microscopy images reveal layer-like and spherical deposits, possibly early stages in a fibrillation process trapped by strong hydrophobic contacts. At higher alcohol concentrations, both DsbB and NhaA are resolubilized and form non-native structures with increased (DsbB) or decreased (NhaA) helicity compared to the native state. The alternative conformational states cannot be returned to the functional native state upon dilution of alcohol. The efficiency of precipitation and the degree to which DsbB is destabilized at low alcohol concentrations show the same correlation with alcohol hydrophobicity. Thus, in addition to their effect on the membrane, alcohols perturb membrane proteins directly by solvating the hydrophobic regions of the protein. At intermediate concentrations, this perturbation exposes hydrophobic segments but does not provide sufficient solvation to avoid intermolecular association. Resolubilization requires a reduction in the relative dielectric constant below 65 in conjunction with specific properties of the individual alcohols. We conclude that alcohols provide access to a diversity of conformations for membrane proteins but are not a priori suitable for solution studies requiring reversible denaturation of monomeric proteins.