Exploring, evolving and exploiting thioester synthetases for industrial biotechnology

goal of biocatalysis is the development of green, enzymatic methods for chemical transformations producing synthetic targets and high-value feedstocks. Current chemical methods for conversion of carboxylic acids to acyl thioesters are complex and multistep. This project aims to develop a one-pot version of this transformation using engineered pimeloyl-CoA synthase (PCAS).
Acyl thioesters are intermediates in the polyketide pathways to many important natural products such as fatty acids, lipids and antibiotics. A recently discovered PCAS from B. subtilis is the key enzyme of vitamin and carboxylation cofactor biotin.
The crystal structure, mechanism and active site residues of this enzyme have been determined and it has been rationally engineered to accept novel fatty acid substrates. This project will fully exploit PCAS and generate enzymes with a broader range of substrates and applications, using directed evolution. A high-throughput screening method will be developed to identify clones with novel activity. Couplings will also be explored which currently lie beyond the reach of synthetic methods.