ENANTIOSELECTIVE SYNTHESIS OF (S)-AMINO ACIDS BY PHENYLALANINE DEHYDROGENASE FROM BACILLUS-SPHAERICUS - USE OF NATURAL AND RECOMBINANT ENZYMES

The substrate specificity of phenylalanine dehydrogenase (L-phenylalanine: NAD+ oxidoreductase, deaminating (EC 1.4.1.-), PheDH) from Bacillus sphaericus SCRC-R79a in the reductive amination reaction was examined with various natural and synthetic 2- and 3-oxo acids and their analogues. The enzyme was active toward 3-substituted pyruvic acids with bulky substituents. Optically pure (S)-phenylalanine and other (S)-amino acids were quantitatively synthesized from their oxo analogues by using PheDH, with a regeneration of NADH by formate dehydrogenase from Candida boidinii No. 2201. Recombinant PheDH overproduced in Escherichia coli JM109/pBPDH1-DBL was effectively used for the syntheses of (S)-amino acids from unnatural substrates. (S)-Phenylalanine could be continuously synthesized for about 1 month with the enzymes, packed in a dialysis tube. It is calculated that 1 molecule of PheDH catalyzed the synthesis of 4.8 x 108 molecules of (S)-phenylalanine, and 2.4 x 105times the weight of (S)-phenylalanine than that of the enzyme. Acetone-dried cells of B. sphaericus SCRC-R79a and C. boidinii No. 2201 were also effective for (S)-phenylalanine synthesis, providing a simple microbial method of synthesis. © 1990, American Chemical Society. All rights reserved.