Zhang Fuzhong, Zarrine-Afsar Arash, Al-Abdul-Wahid M Sameer, Prosser R Scott, Davidson Alan R, Woolley G Andrew
Department of Chemistry, University of Toronto, 80 Saint George Street, Toronto M5S 3H6 Canada.
J Am Chem Soc. 2009 Feb 18;131(6):2283-9. doi: 10.1021/ja807938v.
Photoswitchable proteins offer exciting prospects for remote control of biochemical processes. We propose a general approach to the design of photoswitchable proteins based on the introduction of a photoswitchable intramolecular cross-linker. We chose, as a model, a FynSH3 domain for which the free energy of folding is less than the energy available from photoisomerization of the cross-linker. Taking the experimentally determined structure of the folded protein as a starting point, mutations were made to introduce pairs of Cys residues so that the distance between Cys sulfur atoms matches the ideal length of the cis form, but not the trans form, of the cross-linker. When the trans cross-linker was introduced into this L3C-L29C-T47AFynSH3 mutant, the protein was destabilized so that folded and unfolded forms coexisted. Irradiation of the cross-linker to produce the cis isomer recovered the folded, active state of the protein. This work shows that structure-based introduction of switchable cross-linkers is a feasible approach for photocontrol of folding/unfolding of globular proteins.
光开关蛋白为生化过程的远程控制提供了令人兴奋的前景。我们提出了一种基于引入光开关分子内交联剂来设计光开关蛋白的通用方法。我们选择FynSH3结构域作为模型，其折叠自由能小于交联剂光异构化可获得的能量。以折叠蛋白的实验测定结构为起点，进行突变以引入半胱氨酸残基对，使得半胱氨酸硫原子之间的距离与交联剂顺式形式而非反式形式的理想长度相匹配。当将反式交联剂引入到这个L3C-L29C-T47A FynSH3突变体中时，蛋白质变得不稳定，从而使折叠态和未折叠态共存。交联剂经光照产生顺式异构体后，蛋白质恢复到折叠的活性状态。这项工作表明，基于结构引入可切换交联剂是一种对球状蛋白折叠/去折叠进行光控的可行方法。
