Wroclaw University of Science and Technology, Wroclaw, Poland.
J Chem Theory Comput. 2020 May 12;16(5):3420-3429. doi: 10.1021/acs.jctc.0c00139. Epub 2020 Apr 23.
Currently developed protocols of theozyme design still lead to biocatalysts with much lower catalytic activity than enzymes existing in nature, and, so far, the only avenue of improvement was the in vitro laboratory-directed evolution (LDE) experiments. In this paper, we propose a different strategy based on "reversed" methodology of mutation prediction. Instead of common "top-down" approach, requiring numerous assumptions and vast computational effort, we argue for a "bottom-up" approach that is based on the catalytic fields derived directly from transition state and reactant complex wave functions. This enables direct one-step determination of the general quantitative angular characteristics of optimal catalytic site and simultaneously encompasses both the transition-state stabilization (TSS) and ground-state destabilization (GSD) effects. We further extend the static catalytic field approach by introducing a library of atomic multipoles for amino acid side-chain rotamers, which, together with the catalytic field, allow one to determine the optimal side-chain orientations of charged amino acids constituting the elusive structure of a preorganized catalytic environment. Obtained qualitative agreement with experimental LDE data for Kemp eliminase KE07 mutants validates the proposed procedure, yielding, in addition, a detailed insight into possible dynamic and epistatic effects.
目前开发的拟酶设计方案仍然导致生物催化剂的催化活性远低于自然界中存在的酶，迄今为止，提高催化活性的唯一途径是体外实验室定向进化（LDE）实验。在本文中，我们提出了一种基于“反向”突变预测方法的不同策略。我们反对常见的需要大量假设和大量计算工作的“自上而下”方法，而是主张采用基于直接从过渡态和反应物络合物波函数得出的催化场的“自下而上”方法。这使得能够直接一步确定最佳催化部位的一般定量角特性，同时包含过渡态稳定化（TSS）和基态去稳定化（GSD）效应。我们通过引入氨基酸侧链构象的原子多极子库进一步扩展了静态催化场方法，该库与催化场一起允许确定构成预组织催化环境的难以捉摸结构的带电氨基酸的最佳侧链取向。与 Kemp 消除酶 KE07 突变体的实验 LDE 数据获得定性一致验证了所提出的方法，此外还深入了解了可能的动态和上位效应。
