Simulation-led redesign of polyketide synthase biocatalysts

Enzymes are remarkable biocatalysts that can work under mild conditions and with high specificity and selectivity. If enzymes could be engineered predictably and accurately, they could provide an efficient route to new molecules such as pharmaceuticals. In pharmaceuticals (and many other applications), it is crucially important to control the precise stereochemical structure, as this can make the difference between medicine or poison. In this project, the aim is to develop ways to control how polyketide synthases, an important class of natural biocatalytic machinery, set the stereochemistry of their products, polyketides. This will be done by modifying - or redesigning - a key enzyme in polyketide synthase systems that helps set stereochemistry in the polyketide product chain, a ketoreductase. In particular, in the so-called polyketide synthase type II systems, an acyl-carrier protein (ACP) will bring the evolving polyketide chain to a ketoreductase, which will subsequently set a stereochemical center. Existing structural information helps define how the ACP and the ketoreductase are involved in making the polyketide actinorhodin (a natural antibiotic), and this will guide the development of computational prediction protocols. This will be the bulk of the work, involving protein-protein docking, molecular dynamics simulation and QM/MM reaction simulations. The simulations will predict new ketoreductase variants that alter the stereochemical outcome. To test and improve these computational predictions, experimental characterisation of promising enzyme variants (product outcome, kinetics and structural biology) will be performed. Once successful, the atomic detail of new variants will be confirmed through structural biology techniques (NMR, X-ray crystallography). This interdisciplinary project is thus combining the expertise in computational simulation of enzymes in Bristol and the expertise from an internationally leading academic team with multidisciplinary expertise of polyketide systems and the relevant experimental techniques (enzymology, molecular biology, chemistry and structural biology). Combining simulation and experiment in this way is still developing but will become increasingly important. The strategies and protocols developed in this project, working on key model systems, will therefore be of general use for similar modification of (polyketide) biosynthetic activities.