Sapra K Tanuj, Besir Hüseyin, Oesterhelt Dieter, Muller Daniel J
Max Planck Institute of Molecular Cell Biology and Genetics, 01307 Dresden, Germany.
J Mol Biol. 2006 Jan 27;355(4):640-50. doi: 10.1016/j.jmb.2005.10.080. Epub 2005 Nov 17.
Using single-molecule force spectroscopy we characterized inter- and intramolecular interactions stabilizing structural segments of individual bacteriorhodopsin (BR) molecules assembled into trimers and dimers, and monomers. While the assembly of BR did not vary the location of these structural segments, their intrinsic stability could change up to 70% increasing from monomer to dimer to trimer. Since each stable structural segment established one unfolding barrier, we conclude that the locations of unfolding barriers were determined by intramolecular interactions but that their strengths were strongly influenced by intermolecular interactions. Subtracting the unfolding forces of the BR trimer from that of monomer allowed us to calculate the contribution of inter- and intramolecular interactions to the membrane protein stabilization. Statistical analyses showed that the unfolding pathways of differently assembled BR molecules did not differ in their appearance but in their population. This suggests that in our experiments the membrane protein assembly does not necessarily change the location of unfolding barriers within the protein, but certainly their strengths, and thus alters the probability of a protein to choose certain unfolding pathways.
我们使用单分子力谱法表征了组装成三聚体、二聚体和单体的单个细菌视紫红质（BR）分子结构片段的分子间和分子内相互作用，这些相互作用稳定了这些结构片段。虽然BR的组装并未改变这些结构片段的位置，但其固有稳定性从单体到二聚体再到三聚体可增加高达70%。由于每个稳定的结构片段都建立了一个解折叠屏障，我们得出结论，解折叠屏障的位置由分子内相互作用决定，但其强度受分子间相互作用的强烈影响。从单体的解折叠力中减去BR三聚体的解折叠力，使我们能够计算分子间和分子内相互作用对膜蛋白稳定性的贡献。统计分析表明，不同组装形式的BR分子的解折叠途径在外观上没有差异，但在数量上有差异。这表明在我们的实验中，膜蛋白组装不一定会改变蛋白质内解折叠屏障的位置，但肯定会改变其强度，从而改变蛋白质选择某些解折叠途径的概率。
