Contreras Martínez Lydia M, Borrero Quintana Ernesto E, Escobedo Fernando A, DeLisa Matthew P
School of Chemical and Biomolecular Engineering, Cornell University, Ithaca, New York, USA.
Biophys J. 2008 Mar 1;94(5):1575-88. doi: 10.1529/biophysj.107.119651. Epub 2007 Nov 9.
Protein complementation assays (PCAs) based on split protein fragments have become powerful tools that facilitate the study and engineering of intracellular protein-protein interactions. These assays are based on the observation that a given protein can be split into two inactive fragments and these fragments can reassemble into the original properly folded and functional structure. However, one experimentally observed limitation of PCA systems is that the folding of a protein from its fragments is dramatically slower relative to that of the unsplit parent protein. This is due in part to a poor understanding of how PCA design parameters such as split site position in the primary sequence and size of the resulting fragments contribute to the efficiency of protein reassembly. We used a minimalist on-lattice model to analyze how the dynamics of the reassembly process for two model proteins was affected by the location of the split site. Our results demonstrate that the balanced distribution of the "folding nucleus," a subset of residues that are critical to the formation of the transition state leading to productive folding, between protein fragments is key to their reassembly.
基于分裂蛋白质片段的蛋白质互补分析（PCA）已成为促进细胞内蛋白质-蛋白质相互作用研究与工程设计的强大工具。这些分析基于这样的观察结果：给定的蛋白质可被分裂成两个无活性的片段，且这些片段能重新组装成原始的正确折叠且具有功能的结构。然而，PCA系统在实验中观察到的一个局限性是，相对于未分裂的亲本蛋白质，蛋白质从其片段折叠的速度要慢得多。部分原因是对PCA设计参数（如一级序列中的分裂位点位置和所得片段的大小）如何影响蛋白质重新组装效率的理解不足。我们使用了一个极简的晶格模型来分析两个模型蛋白质重新组装过程的动力学如何受到分裂位点位置的影响。我们的结果表明，“折叠核”（对导致有效折叠的过渡态形成至关重要的一部分残基）在蛋白质片段之间的平衡分布是它们重新组装的关键。
