Department of Molecular Genetics, the University of Toronto, Toronto, Ontario M5S 1A8, Canada.
Science. 2010 Sep 10;329(5997):1312-6. doi: 10.1126/science.1191723.
Proteins can sample conformational states that are critical for function but are seldom detected directly because of their low occupancies and short lifetimes. In this work, we used chemical shifts and bond-vector orientation constraints obtained from nuclear magnetic resonance relaxation dispersion spectroscopy, in concert with a chemical shift-based method for structure elucidation, to determine an atomic-resolution structure of an "invisible" folding intermediate of a small protein module: the FF domain. The structure reveals non-native elements preventing formation of the native conformation in the carboxyl-terminal part of the protein. This is consistent with the kinetics of folding in which a well-structured intermediate forms rapidly and then rearranges slowly to the native state. The approach introduces a general strategy for structure determination of low-populated and transiently formed protein states.
蛋白质可以探测对其功能至关重要的构象状态，但由于这些状态的占有率低、寿命短，很少能够直接检测到。在这项工作中，我们使用核磁共振弛豫弥散光谱获得的化学位移和键矢量方向约束，结合基于化学位移的结构解析方法，确定了一个小蛋白模块的“不可见”折叠中间体的原子分辨率结构：FF 结构域。该结构揭示了阻止蛋白质羧基末端形成天然构象的非天然元件。这与折叠动力学一致，其中一个结构良好的中间体快速形成，然后缓慢重排到天然状态。该方法为低占有率和瞬时形成的蛋白质状态的结构测定引入了一种通用策略。
