Asymmetrical Synthesis of l-Homophenylalanine Using Engineered Escherichia coli Aspartate Aminotransferase

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Abstract
Site‐directed mutagenesis was performed to change the substrate specificity of Escherichia coli aspartate aminotransferase (AAT). A double mutant, R292E/L18H, with a 12.9‐fold increase in the specific activity toward l‐lysine and 2‐oxo‐4‐phenylbutanoic acid (OPBA) was identified. E. coli cells expressing this mutant enzyme could convert OPBA to l‐homophenylalanine (l‐HPA) with 97% yield and more than 99.9% ee using l‐lysine as amino donor. The transamination product of l‐lysine, 2‐keto‐6‐aminocaproate, was cyclized nonenzymatically to form Δ1‐piperideine 2‐carboxylic acid in the reaction mixture. The low solubility of l‐HPA and spontaneous cyclization of 2‐keto‐6‐aminocaproate drove the reaction completely toward l‐HPA production. This is the first aminotransferase process using l‐lysine as inexpensive amino donor for the l‐HPA production to be reported.

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