Chaturvedi Shobhit S, Vargas Santiago, Ajmera Pujan, Alexandrova Anastassia N
Department of Chemistry and Biochemistry, University of California, Los Angeles, California 90095, United States.
J Am Chem Soc. 2024 Jun 7. doi: 10.1021/jacs.4c03914.
To unravel why computational design fails in creating viable enzymes, while directed evolution (DE) succeeds, our research delves into the laboratory evolution of protoglobin. DE has adapted this protein to efficiently catalyze carbene transfer reactions. We show that the previously proposed enhanced substrate access and binding alone cannot account for increased yields during DE. The 3D electric field in the entire active site is tracked through protein dynamics, clustered using the affinity propagation algorithm, and subjected to principal component analysis. This analysis reveals notable changes in the electric field with DE, where distinct field topologies influence transition state energetics and mechanism. A chemically meaningful field component emerges and takes the lead during DE and facilitates crossing the barrier to carbene transfer. Our findings underscore intrinsic electric field dynamic's influence on enzyme function, the ability of the field to switch mechanisms within the same protein, and the crucial role of the field in enzyme design.
为了揭示为什么计算设计在创建可行的酶时失败，而定向进化（DE）却成功，我们的研究深入探讨了原球蛋白的实验室进化。DE已使这种蛋白质适应于高效催化卡宾转移反应。我们表明，先前提出的仅增强底物可及性和结合不能解释DE过程中产量的增加。通过蛋白质动力学跟踪整个活性位点的三维电场，使用亲和传播算法进行聚类，并进行主成分分析。该分析揭示了DE过程中电场的显著变化，其中不同的场拓扑结构影响过渡态能量和机制。一个具有化学意义的场成分出现并在DE过程中起主导作用，促进跨越卡宾转移的障碍。我们的发现强调了内在电场动力学对酶功能的影响、场在同一蛋白质内切换机制的能力以及场在酶设计中的关键作用。
