Combinations of affinity-enhancing mutations in a T cell receptor reveal highly nonadditive effects within and between complementarity determining regions and chains.
Combinations of affinity-enhancing mutations in a T cell receptor reveal highly nonadditive effects within and between complementarity determining regions and chains.
复制标题
T 细胞受体中亲和力增强突变的组合揭示了互补决定区和链内部和之间的高度非加和效应。
DOI：
10.1021/bi901969a
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发表时间：
2010
期刊：
影响因子：
2.9
通讯作者：
Weng,Zhiping
中科院分区：
文献类型：
--
作者：
Pierce,BrianG;Haidar,JaafarN;Yu,Yong;Weng,Zhiping
理解蛋白质-蛋白质界面中对多个突变的能量和结构响应是合理蛋白质设计的一个关键方面。在这里，我们调查的T细胞受体（TCR），bindin vivo HLA-A2 MHC和病毒肽的点突变的组合的协同性。这些突变来自两个来源：TCR α链的基于结构的设计研究（9个突变）和TCR β链的体外选择研究（4个突变）。除了结合来自每条链的最高亲和力变体之外，我们还测试了链内和链间的其他突变组合，总共有23种TCR突变体，我们测量了它们与肽和主要组织相容性复合体的结合动力学。观察到广泛的结合亲和力，与野生型相比，从2倍到1000倍的结合改善，在TCR链内和之间观察到显著的非累加效应。这包括在野生型复合物中相隔超过9个碱基的CDR 1和CDR 3残基之间的氨基酸依赖性协同相互作用。当分析突变的动力学时，我们发现缔合速率主要负责协同性，而解离速率负责反协同性（小于加性能量学）。反合作突变体的结构建模的基础上，我们确定，近端突变体之间的侧链冲突可能导致非加性结合能。这些结果突出了TCR缔合和结合的复杂性质，并将在未来的设计工作中提供信息，联合收割机组合多个突变残基。
Understanding the energetic and structural response to multiple mutations in a protein−protein interface is a key aspect of rational protein design. Here we investigate the cooperativity of combinations of point mutations of a T cell receptor (TCR) that bindsin vivoto HLA-A2 MHC and a viral peptide. The mutations were obtained from two sources: a structure-based design study on the TCR α chain (nine mutations) and anin vitroselection study on the TCR β chain (four mutations). In addition to combining the highest-affinity variants from each chain, we tested other combinations of mutations within and among the chains, for a total of 23 TCR mutants that we measured for binding kinetics to the peptide and major histocompatibility complex. A wide range of binding affinities was observed, from 2- to 1000-fold binding improvement versus that of the wild type, with significant nonadditive effects observed within and between TCR chains. This included an amino acid-dependent cooperative interaction between CDR1 and CDR3 residues that are separated by more than 9 Å in the wild-type complex. When analyzing the kinetics of the mutations, we found that the association rates were primarily responsible for the cooperativity, while the dissociation rates were responsible for the anticooperativity (less-than-additive energetics). On the basis of structural modeling of anticooperative mutants, we determined that side chain clash between proximal mutants likely led to nonadditive binding energies. These results highlight the complex nature of TCR association and binding and will be informative in future design efforts that combine multiple mutant residues.