Whole-cell biocatalytic production of variously substituted β-aryl- and β-heteroaryl-β-amino acids

Abstract

Biologically-active β-peptides and pharmaceuticals that contain key β-amino acids are emerging as target therapeutics; thus, synthetic strategies to make substituted, enantiopure β-amino acids are increasing. Here, we use whole-cell Escherichia coli (OD600 ∼35) engineered to express a Pantoea agglomerans phenylalanine aminomutase (PaPAM) biocatalyst. In either 5mL, 100mL, or 1L of M9 minimal medium containing α-phenylalanine (20mM), the cells produced ∼1.4mgmL−1 of β-phenylalanine in each volume. Representative pilot-scale 5-mL cultures, fermentation reactions converted 18 variously substituted α-arylalanines to their (S)-β-aryl-β-amino acids in vivo and were not toxic to cells at mid- to late-stage growth. The β-aryl-β-amino acids made ranged from 0.043mg (p-nitro-β-phenylalanine, 4% converted yield) to 1.2mg (m-bromo-β-phenylalanine, 96% converted yield) over 6h in 5mL. The substituted β-amino acids made herein can be used in redox and Stille-coupling reactions to make synthetic building blocks, or as bioisosteres in drug design.