Ultrahigh-throughput protein evolution for polyethylene biodegradation

ended

The build-up of plastic pollution is one of the most pressing environmental concerns. Polyethylene (PE), the most abundantly produced plastic polymer, can persist in nature for over a century. The microbial biodegradation of PE that has been observed is slow and inefficient. So far no effort has been undertaken to improve the efficiency of enzymes involved in the biodegradation of PE through directed protein evolution. Standard assays for measuring degradation rates are not sufficiently high-throughput to cover the sequence space required. I propose to use state-of-the-art protein evolution technology to overcome this problem in two ways. First, an ultrahigh-throughput microfluidics based approach, that can associate a given genotype with its phenotype in picoliter sized water-in-oil droplets, will be used to isolate the desired genotypes from a random mutagenesis library. Second, a novel assay for measuring polymer concentration within each droplet based on differential light scattering as the polymer is degraded will assay the PE degradation rate for a given enzyme. These techniques were developed in the research group of the proposed host, Dr. Hollfelder in the Department of Biochemistry at the University of Cambridge. Using these techniques, I will functionally express and evolutionarily optimise a range of PE degrading enzymes in genetically tractable host strains, creating a chassis to investigate the potential of microbial biodegradation as a solution to plastic waste. Secondments at the EBI, UCL and the SME Drop-Tech will convey practical skills in bioinformatics screens and droplet formation. The host group’s experience in enzyme biotechnology and directed protein evolution as well as its extensive modern facilities for microfluidics, next generation sequencing and flow cytometry will synergize with my personal research experience in synthetic, molecular and microbiology to find a multidisciplinary solution to the growing problem of plastic degradation.