Abstract
Enzymes are responsible for many chemical reactions which support life and are used in numerous industries. Laboratory evolution experiments can mimic the Darwinian process of mutation and selection pressure, yielding newly evolved catalysts. Recently developed single-cell microencapsulation techniques enable the screening of ultralarge enzyme-mutant pools, greatly increasing the odds of isolating improved catalysts. The encapsulation process uses droplet microfluidics, a groundbreaking technology for the miniaturization and automation of biochemical assays which has already demonstrated its effectiveness at evolving enzymes and identifying ultrarare catalysts from the environment. This chapter reviews how large-scale single-cell assays provide an efficient route toward the identification of biocatalysts with novel or improved function. Single-cell microencapsulation enables retention of genotype (gene sequence) and phenotype (enzyme function) within a stable microcompartment. In practice, single-gene mutants expressed by single cells with their corresponding enzyme are encapsulated into monodisperse water-in-oil microdroplets where the assay takes place. This enables the screening of ultralarge 108 members libraries in a day with a concomitant reduction of 1000-fold in assay-reagent costs and a 1000-fold increase in throughput compared to conventional tools. This technology has already been widely adopted for the accelerated discovery and evolution of biocatalysts used in many application domains from the decontamination of polluted environments, bioenergy production, to the degradation of plastics and the discovery of antimicrobials.
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Gielen, F. (2022). Single-Cell Microencapsulation for Evolution and Discovery of Biocatalysts. In: Santra, T.S., Tseng, FG. (eds) Handbook of Single-Cell Technologies. Springer, Singapore. https://doi.org/10.1007/978-981-10-8953-4_47
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