Department of Agricultural Biotechnology and Research Institute of Agriculture and Life Sciences, Seoul National University, 1 Gwanak-ro, Gwanak-gu, Seoul 08826, Republic of Korea.
Protein Structure Research Team, Korea Basic Science Institute, 162 Yeongudanji-Ro, Ochang, Chungbuk 28119, Republic of Korea.
Structure. 2018 Jun 5;26(6):887-893.e2. doi: 10.1016/j.str.2018.04.009. Epub 2018 May 17.
Proteins assemble to form functional complexes via the progressive evolution of nonspecific complexes formed by transient encounters. This target search process generally involves multiple routes that lead the initial encounters to the final complex. In this study, we have employed NMR paramagnetic relaxation enhancement to visualize the encounter complexes between histidine-containing phosphocarrier protein and the N-terminal domain of enzyme I and demonstrate that protein association can be significantly enhanced by engineering on-pathways. Specifically, mutations in surface charges away from the binding interface can elicit new on-pathway encounter complexes, increasing their binding affinity by an order of magnitude. The structure of these encounter complexes indicates that such on-pathways extend the built-in target search process of the native protein complex. Furthermore, blocking on-pathways by countering mutations reverts their binding affinity. Our study thus illustrates that protein interactions can be engineered by rewiring the target search process.
蛋白质通过非特异性复合物的渐进进化形成功能性复合物，这些非特异性复合物是由短暂的相遇形成的。这个目标搜索过程通常涉及多个途径，这些途径将最初的相遇引导到最终的复合物。在这项研究中，我们利用 NMR 顺磁弛豫增强来可视化含组氨酸的磷酸载体蛋白与酶 I N 端结构域之间的相遇复合物，并证明通过工程途径可以显著增强蛋白质的缔合。具体来说，远离结合界面的表面电荷突变可以诱发生成新的通路上的相遇复合物，将它们的结合亲和力提高一个数量级。这些相遇复合物的结构表明，这种通路上的延伸扩展了天然蛋白质复合物的内置目标搜索过程。此外，通过抵消突变来阻断通路上的作用会使其结合亲和力恢复正常。因此，我们的研究表明，可以通过重新布线目标搜索过程来设计蛋白质相互作用。
