Biocatalytic synthesis of N-methyl-L-phenylalanine by phenylalanine ammonia lyase from galanthamine-producing Lycoris radiata

Abstract:

[Background] N-Methyl-L-phenylalanine, an N-alkylated aromatic amino acid, is a valuable chiral building block/intermediate/ingredient presented in many specialized metabolites that are very important in pharmaceutical, nutraceutical, and agrochemical industries. The synthesis and preparation of N-alkylated aromatic amino acids from aromatic α,β-unsaturated carboxylic acid remain challenging. [Objective] Herein we report a one-step biologically catalyzed N-methylamination of trans-cinnamic acid by LrPAL3, a phenylalanine ammonia lyase (PAL) from galanthamine-producing Lycoris radiata, to generate N-methyl-L-phenylalanine. [Methods] HPLC-DAD and HRESIMS analyses revealed that N-methyl-phenylalanine was produced when incubated trans-cinnamic acid and methylamine using the whole Escherichia coli BL21(DE3) cells expressing LrPAL3 as catalyst. [Results] The ¹H-NMR data and optical rotation of the enzymatic bioconversion product are in agreement with those of the authentic N-methyl-L-phenylalanine, which demonstrated the LrPAL3-catalyzed one-step regio- and enantioselective N-methylamination product of trans-cinnamic acid is N-methyl-L-phenylalanine. [Conclusion] This work provides an alternative biocatalyst for the asymmetric synthesis of valuable chiral N-methyl-L-phenylalanine. It paves a way to biologically synthesize N-alkylated amino acids through metabolic engineering and direct protein evolution of LrPAL3.