Simulation-led engineering of Diels-Alderase activity and selectivity for sustainable biosynthesis of new antibiotics

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Enzymes are remarkable biocatalysts that allow rapid, selective and efficient catalysis under mild conditions.Biotechnology exploits the rate-enhancing properties of these natural biocatalysts to manufacture high added-valueproducts. Of particular current interest are enzymes that catalyse the Diels-Alder reaction, a [4+2] cycloaddition reactionwidely recognised as one of the cornerstone synthetic organic reactions of the 20th century. It is commonly employed inthe synthesis of bioactive natural products, including numerous important pharmaceuticals. Without enzyme biocatalysts,harsh reaction conditions (high temperature, high pressure) often must be employed, and precise control over the productoutcome is limited. Enzymes that catalyse this reaction, Diels-Alderases, are therefore highly attractive. In this project, the focus will be on Diels-Alderases involved in the biosynthesis of tetronate antibiotics such asabyssomicins, natural products with highly promising antibiotic activity. We have already established the structure andmechanism of two of these in detail, for which computational simulation, including docking, molecular dynamicsand QM/MM reaction simulations, has proven highly valuable. The same is true for crucial tailoring enzymes that arerequired before and after the Diels-Alderase step to arrive at the active polyketide-based antibiotic. The project aims touse these techniques in new computational prediction protocols that can suggest enzyme variants with desired changesin activity and specifity, for example to obtain tetronate antiobiotics with improved characteristics for pharmaceuticaluse. To test and improve these predictions, experimental characterisation of promising enzyme variants (productoutcome, kinetics and structural biology) will be performed. By working with our industrial partner AstraZeneca, theimpact of engineered enzymes can be realised by testing their use for generating valuable building blocks and scaffolds forpharmaceutical drug discovery. This interdisciplinary project combines the expertise in computational simulation ofenzymes in Bristol and the expertise from an internationally leading academic team with multidisciplinary expertise ofpolyketide natural product biosynthesis and 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; in Bristol, weare at the forefront of this development. The multidisciplinary environment ensures the student will acquire a range ofskills that will arm them for a future career in academic or industrial bioscience (including pharmaceutical science). Thestudent will be embedded in the vibrant research environment in Bristol, including the Centre for ComputationalChemistry and the Bristol BioDesign institute, ensuring a wide range of interactions, seminar programmes and courses.