Evolving a stereochemically promiscuous aldolase to create new classes of stereoselective biocatalyst for asymmetric synthesis

该提案描述了使用热稳定酶来制备新型手性化合物，作为用于合成制药和精细化工工业的重要中间体的结构单元。将研究酶催化与广泛底物的特定反应的范围和限制。该酶含有一个催化发生的活性位点，其结构已通过X射线晶体学在分子水平上表征。这个活性位点的结构的知识将使我们能够产生新的“突变体”催化剂与自己的特异性配置文件作为改进的生物催化剂的合成。这些突变研究将分为两类：首先，我们将故意将特定的变化引入酶中;其次，我们将产生随机突变，以提供大量的催化剂库。筛选协议将开发，这将使我们能够快速识别具有有用的催化活性的突变体，目的是识别进行目前在自然界中未观察到的反应的生物催化剂。同时，我们还将开发一种化学方法来合成这些类型的化合物，使用小手性分子作为替代催化剂。本研究中产生的许多化合物将转化为用于合成具有生物重要性的天然产物的新型高度功能化的中间体。

This proposal describes the use of a thermostable enzyme to prepare new types of chiral compounds as building blocks for synthesising important intermediates for the pharmaceutical and fine chemical industry. The scope and limiation of the enzyme to catalyse a specific reaction with a wide range of substrates will be investigated. The enzyme contains an active site where catalysis takes place, and its structure has been characterised at a molecular level by X-ray crystallography. Knowledge of the structure of this active site will enable us to generate new 'mutant' catalysts with their own specificity profiles as improved biocatalysts for synthesis. These mutation studies will fall into two categories: firstly, we will deliberately introduce specific changes into the enzyme; and secondly, we will generate random mutations to afford large libraries of catalysts. Screening protocols will be developed that will enable us to rapidly identify mutants with useful catalytic activity, with the aim of identifying biocatalysts that carry out reactions not currently observed in nature. Concurrently, we will also develop a chemical approach to the synthesis of these type of compounds, using small chiral molecules as alternative catalysts. A number of compounds produced in this study will then be converted into novel highly fuctionalised intermediates of use for the synthesis of biologically-important natural products.