Of the enzymes evaluated, ATA TA25 (previously reported as 3FCR-QM-W59L) gave the highest conversions of 6 and 7 to the respective amine products (Figure ). To unlock desired aminotransferase activity with ketone 5, we subjected TA25 to four rounds of directed evolution using a substrate walking approach (Figure and Supplementary Table ). In each round, ca. 20-24 positions were individually randomized using NNK degenerate codons. Individual library variants were arrayed in 96-well plates and evaluated as clarified cell lysate using UPLC assays. Beneficial mutations identified in each round were subsequently combined by DNA shuffling.