Genome scrambling and biosensor technologies for production of high value chemicals in synthetic yeast (Chem@Sc2.0)

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The yeast synthetic genome project, the largest Synthetic Biology project to date, provides yeast, Saccharomyces cerevisiae 2.0 (Sc2.0), with chromosomal loxP sites, enabling Synthetic Chromosome Recombination and Modification by LoxP-mediated Evolution (SCRaMbLE, Science 2017, 355, 1040-1045). In this project, we will exploit Sc2.0 as a platform for producing valuable natural products (NP), including essential antibiotics (e.g. Nature Catalysis 2018, 1, 977-984), anticancer agents, immunosuppressants and statins (blockbuster drugs). Unlike the wild-type S. cerevisiae, Sc2.0 has synthetic chromosomes (re-designed DNA sequences chemically synthesized), enabling the genome to be scrambled, generating mutants where genes have been rearranged, deleted or duplicated. We have shown that, by introducing heterologous biosynthetic gene clusters into Sc2.0, it is possible to generate scrambled mutants with enhanced capacity for producing target compounds (Nature Commun. 2018, 9, 1936). However, screening large numbers (billions) of Sc2.0 mutants generated by genome scrambling is challenging. To this end, we will develop biosensors, based on modular riboswitch components (e.g. J. Am. Chem. Soc. 2015, 137, 9015-9021 & J. Am. Chem. Soc. 2014, 136, 10615-10624), that can bind to the target NP and trigger a fluorescent response. By introducing biosensors into Sc2.0, along with the genes required for the biosynthesis of the target NP, we will be able to rapidly select the scrambled mutant cells that produce the highest levels of the target compound; these cells will be fluorescent and can be easily separated using a fluorescence activated cell sorter. Strains producing highest yields of desired compounds, will be subjected to further rounds of scrambling with biosensor screening, and their genomes will be sequenced. The project will provide a paradigm shift in pathway (metabolic) engineering, that could lead to more cost-effective and sustainable production of antibiotics required to combat antimicrobial resistance (AMR), as well as other essential medicines. This ambitious project provides cutting-edge training in synthetic biology, spanning NP biosynthesis, RNA-based biosensors and genetics. The project will be supervised by Profs Jason Micklefield and Patrick Cai, and will be based in the Manchester Institute of Biotechnology (MIB) and SYNBIOCHEM centre at the UoM, with world-class facilities and training opportunities.