Sequence-dependent folding of single proteins under mechanical load

蛋白质必须折叠成精确的三维结构才能正常发挥作用。折叠过程是细胞内遗传信息流从存储到有用部署的重要组成部分。尽管对蛋白质折叠的研究已经有了几十年的历史，但由于直接测量折叠转变的实验挑战，关于氨基酸序列如何影响折叠仍有许多需要了解的地方。这个项目将探索折叠的基本原理，通过使用光学陷阱施加的机械力来重复展开和重新折叠单个蛋白质分子。通过监测平衡(恒定力)和非平衡(变化力)条件下的分子端到端距离，将以高精度测量单个折叠轨迹。根据这些轨迹将确定折叠过程的定量热力学和动力学性质，包括折叠的自由能、折叠和去折叠速率随力的函数、中间态在反应坐标上的存在和位置以及势垒的位置和高度。通过改变简单蛋白质结构域的氨基酸序列，系统地研究了序列对这些性质的影响。

Proteins must fold into precise three-dimensional structures in order to function correctly. The folding process is an essential part of the flow of genetic information in the cell from storage to useful deployment. Despite many decades of research on protein folding, much remains to be learned about how the amino acid sequence affects folding, due to the experimental challenges in measuring the folding transition directly. This project will probe the basic principles of folding by using mechanical force applied by an optical trap to unfold and refold single protein molecules repeatedly. Individual folding trajectories will be measured with high-precision by monitoring the molecular end-to-end distance under both equilibrium (constant force) and non-equilibrium (changing force) conditions. Quantitative thermodynamic and kinetic properties of the folding process will be determined from these trajectories, including the free energy for folding, the folding and unfolding rates as a function of force, the presence and location along the reaction coordinate of intermediate states, and the location and height of energy barriers. The effect of sequence on these properties will be investigated systematically by varying the amino acid sequence of simple protein domains.