Fig. 4: Directed evolution of SFC_V15pAPhC for the enantioselective Friedel-Crafts alkylation reaction.
From: Design and evolution of artificial enzyme with in-situ biosynthesized non-canonical amino acid
a Asymmetric Friedel-Crafts alkylation of enal 1a and indole 2a to form alkylation product (R)−3a by SFC_V15pAPhC mutants. b Enhanced view of the hydrophobic pocket showing selected positions for enzyme directed evolution. c The iterative-site-specific mutagenesis strategy for the direct evolution of SFC_V15pAPhC. d Reaction outcomes of e.e. (yellow) and yield (blue) afforded by different evolved SFC_V15pAPhC variants. Reaction conditions: Enzyme varaints (5.0 mol% dimer concentration), 5 mM 1a and 1 mM 2a in 50 mM NaH2PO4, 150 mM NaCl (pH 6.5) buffer with DMF (4.0% v/v). Reactions conducted for 10 h at 4 °C and analyzed by normal-phase chiral column with e.e. and yield, unless noted otherwise. Yields are determined by HPLC analysis using a calibration curve of pure product 3a. Error bars represent ± standard deviation of the mean over 3 independent replicates. Source data are provided as a Source Data file.
