ArticleAnalytical chemistry2025
A Microfluidics-Based Ultrahigh-Throughput Screening Unveils Diverse Ketoreductases Relevant to Pharmaceutical Synthesis.
Article in Analytical chemistry, 2025. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Not yet cited in PubMed.
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Abstract
Ketoreductases (KREDs) have become increasingly valuable biocatalysts due to their ability to produce chiral alcohols with high enantioselectivity. Prior to our work, Thai et al. developed an efficient and easy assay for their discovery, but the throughput was limited. Based on their work, we developed an ultrahigh-throughput screening assay to discover KREDs. First, we optimized Thai's assay by adapting it to a droplet format and increased its throughput by combining droplet microfluidics and fluorescence-activated cell sorting (FACS). Then, we demonstrated that our new assay was reliable and sensitive by successfully screening a library of 1.5 million clones. This allowed us to discover KREDs with low identity with known enzymes or with a previously undescribed substrate scope, which could not have been predicted computationally. In conclusion, our assay was used to carry out the first metagenomic screening for KREDs in microdroplets, and it can be used to screen any large KRED library toward enzyme discovery or evolution, as well as to enable coupled ultrahigh-throughput screening assays for other enzyme activities.
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