Ultrahigh-throughput FACS-based screening for directed enzyme evolution.
Directed enzyme evolution has proven to be a powerful tool for improving a range of properties of enzymes through consecutive rounds of diversification and selection. However, its success depends heavily on the efficiency of the screening strategy employed. Fluorescence-activated cell sorting (FACS) has recently emerged as a powerful tool for screening enzyme libraries due to its high sensitivity and its ability to analyze as many as 10(8) mutants per day. Applications of FACS screening have allowed the isolation of enzyme variants with significantly improved activities, altered substrate specificities, or even novel functions. This review discusses FACS-based screening for enzymatic activity and its potential application for the directed evolution of enzymes, ribozymes, and catalytic antibodies.
Yang
Guangyu
G
Centre for High-Throughput Biology (CHiBi) and Department of Chemistry, University of British Columbia, 2036 Main Mall, Vancouver, B.C. V6T 1Z1, Canada.
Withers
Stephen G
SG
D000595:Amino Acid Sequence; D000818:Animals; D019020:Directed Molecular Evolution / Q000379:methods*; D004798:Enzymes / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D005434:Flow Cytometry / Q000295:instrumentation / Q000379:methods*; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000737:chemistry / Q000378:metabolism; D057166:High-Throughput Screening Assays / Q000295:instrumentation / Q000379:methods*; D015394:Molecular Structure; D015202:Protein Engineering / Q000295:instrumentation / Q000379:methods; D013379:Substrate Specificity
D004798:Enzymes; D049452:Green Fluorescent Proteins