Effcient identifcation and screening of functional enzyme libraries for functional enzyme resource mining

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Saccharomyces cerevisiae FlowRACS

Highlights

• With single-cell Raman spectroscopy, characteristic peaks correlated with diacylglycerol acyltransferase activity can be identified

• Rapid screening of functional enzyme libraries can be accomplished on FlowRACS®, High-throughput Raman-activated Flow Cytometer

• With FlowRACS®, 3 previously reported strong genes and 2 novel weak genes were obtained in a 10-minute run,compared to months of work using conventional approaches

Abstract

Diacylglycerol acyltransferase (DGAT) is a key enzyme catalyzing the final step of lipid synthesis in human, animals, and plants, and is the only rate-limiting enzyme. Using FlowRACS®, label-free,high-throughput, high-accuracy, non-destructive sorting of DGAT mutant libraries can be achieved at single-cell resolution. In this experiment, from a library of candidate DGAT genes derived from Micrococcus, three previously reported strong genes and two never-reported weak genes were obtained by FlowRACS® in a10-minute run, whereas the screening and characterization of these three strong functional genes with traditional methods would have taken several months. In conclusion, FlowRACS® provides a powerful tool for mining highly efficient enzymes.

Graphical Abstract

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