Thomas M. Groseclose, Erin A. Kober, Matilda Clark, Benjamin Moore, Shounak Banerjee, Victoria Bemmer, Gregg T. Beckham, Andrew R. Pickford, Taraka T. Dale, Hau B. Nguyen
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引用次数: 0
Abstract
The ability of enzymes to hydrolyze the ubiquitous polyester, poly(ethylene terephthalate) (PET), has enabled the potential for bioindustrial recycling of this waste plastic. To date, many of these PET hydrolases have been engineered for improved catalytic activity and stability, but current screening methods have limitations in screening large libraries, including under high-temperature conditions. Here, we developed a platform that can simultaneously interrogate PET hydrolase libraries of 104–105 variants (per round) for protein solubility, thermostability, and activity via paired, plate-based split green fluorescent protein and model substrate screens. We then applied this platform to improve the performance of a benchmark PET hydrolase, leaf-branch compost cutinase, by directed evolution. Our engineered enzyme exhibited higher catalytic activity relative to the benchmark, LCC-ICCG, on amorphous PET film coupon substrates (∼9.4% crystallinity) in pH-controlled bioreactors at both 65 °C (8.5% higher conversion at 48 h and 38% higher maximum rate, at 2.9% substrate loading) and 68 °C (11.2% higher conversion at 48 h and 43% higher maximum rate, at 16.5% substrate loading), up to 48 h, highlighting the potential of this screening platform to accelerate enzyme development for PET recycling.
期刊介绍:
ACS Catalysis is an esteemed journal that publishes original research in the fields of heterogeneous catalysis, molecular catalysis, and biocatalysis. It offers broad coverage across diverse areas such as life sciences, organometallics and synthesis, photochemistry and electrochemistry, drug discovery and synthesis, materials science, environmental protection, polymer discovery and synthesis, and energy and fuels.
The scope of the journal is to showcase innovative work in various aspects of catalysis. This includes new reactions and novel synthetic approaches utilizing known catalysts, the discovery or modification of new catalysts, elucidation of catalytic mechanisms through cutting-edge investigations, practical enhancements of existing processes, as well as conceptual advances in the field. Contributions to ACS Catalysis can encompass both experimental and theoretical research focused on catalytic molecules, macromolecules, and materials that exhibit catalytic turnover.