Department of Chemistry, University of Toronto, 80 Saint George Street, Toronto, Ontario M5S 3H6, Canada.
Biochemistry. 2012 Aug 14;51(32):6421-31. doi: 10.1021/bi300685a. Epub 2012 Aug 3.
Photoswitchable distance constraints in the form of photoisomerizable chemical cross-links offer a general approach to the design of reversibly photocontrolled proteins. To apply these effectively, however, one must have guidelines for the choice of cross-linker structure and cross-linker attachment sites. Here we investigate the effects of varying cross-linker structure on the photocontrol of folding of the Fyn SH3 domain, a well-studied model protein. We develop a theoretical framework based on an explicit-chain model of protein folding, modified to include detailed model linkers, that allows prediction of the effect of a given linker on the free energy of folding of a protein. Using this framework, we were able to quantitatively explain the experimental result that a longer, but somewhat flexible, cross-linker is less destabilizing to the folded state than a shorter more rigid cross-linker. The models also suggest how misfolded states may be generated by cross-linking, providing a rationale for altered dynamics seen in nuclear magnetic resonance analyses of these proteins. The theoretical framework is readily portable to any protein of known folded state structure and thus can be used to guide the design of photoswitchable proteins generally.
以光致异构化学交联形式存在的光可切换距离约束为设计可还原光控蛋白质提供了一种通用方法。然而，要有效地应用这些方法，必须有关于交联剂结构和交联剂附着位点选择的指导方针。在这里，我们研究了交联剂结构变化对 Fyn SH3 结构域折叠的光控的影响，Fyn SH3 结构域是一种研究得很好的模型蛋白。我们开发了一个基于蛋白质折叠的显链模型的理论框架，该模型经过修改，包括详细的模型接头，允许预测给定接头对蛋白质折叠自由能的影响。使用这个框架，我们能够定量解释实验结果，即较长但有些灵活的交联剂对折叠状态的稳定性不如较短更刚性的交联剂。该模型还表明交联如何产生错误折叠状态，为这些蛋白质的核磁共振分析中观察到的动力学变化提供了合理的依据。该理论框架易于移植到任何具有已知折叠状态结构的蛋白质，因此可用于指导一般的光可控蛋白质的设计。
