A Pantoea agglomerans-derived phenylalanine aminomutase (PaPAM) was engineered to improve the biocatalytic synthesis of (S)-beta-phenylalanine, which is an important precursor of pharmaceuticals and peptidomimetics. A semi-rational design strategy based on a combination of surface-amino-acid engineering and the amino acid preference of the thermozyme was applied to counteract the enzyme tradeoff between improving its activity and stability. The surface glycine, lysine and serine of PaPAM were mutated to alanine, arginine and alanine, respectively. A K340R mutant was screened with a 2.23-fold increased activity and 2.12-fold improved half-life at 50 degrees C over those of the wild-type PaPAM. These improvements resulted from the more stable enzymatic conformation as well as the more rigid inner loop in K340R. When tested in a whole-cell biocatalytic reaction, the (S)-beta-phenylalanine volumetric productivity of K340R reached 0.47 g/L.h (1.4-fold greater than that of the wild-type PaPAM), and the conversion rate was improved by 17% compared to that of the wild-type PaPAM. The enzymatic properties of K340R and the resulting (S)-beta-phenylalanine production are among the highest reported, and the results indicate that the described strategy is potent for engineering enzymatic stability and activity of PAM. (C) 2019 Elsevier Inc. All rights reserved.