Asymmetric synthesis of chiral amines with omega-transaminase.
The asymmetric synthesis of chiral amines using prochiral ketones was carried out with (S)-specific omega-transaminase (omega-TA) from Vibrio fluvialis JS17. This reaction is inhibited severely by both products, (S)-amine and deaminated ketone. In addition, thermodynamic equilibrium strongly favored the reverse reaction. L-Alanine proved to be the best amino donor based on easy removal of the products. Optimal pH of the reactions with both whole cells and cell-free extract was 7. Amino acceptor reactivities of ketone substrates and reaction profiles of the asymmetric synthesis showed that the initial rate as well as the reaction yield were lower when the resulting (S)-amine from a prochiral ketone substrate was a more reactive amino donor. The yield could be increased dramatically by removing pyruvate, which is a more inhibitory product than (S)-alpha-methylbenzylamine [(S)-alpha-MBA] when acetophenone and L-alanine are used as an amino acceptor and donor, respectively. The removal of pyruvate was carried out by incorporating lactate dehydrogenase (LDH) in cell-free extract or by using whole cells. The whole cell reaction yielded a much better result. When 25 mM benzylacetone and 30 mM acetophenone were used as an amino acceptor with 300 mM L-alanine, 90.2% and 92.1% of the reaction yields after 1 day were obtained with whole cells, respectively. Enantiomeric excesses of both (S)-alpha-MBA and (S)-1-methyl-3-phenylpropylamine [(S)-MPPA] were all above 99%.
Shin
J S
JS
Laboratory of Molecular Biotechnology and Biomaterials, Institute for Molecular Biology and Genetics, Seoul National University, Seoul 151-742, Korea.
Kim
B G
BG
D000409:Alanine / Q000378:metabolism; D000588:Amines / Q000138:chemical synthesis*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007659:Ketones / Q000378:metabolism / Q000494:pharmacology; D007700:Kinetics; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008968:Molecular Conformation; D019289:Pyruvic Acid / Q000494:pharmacology; D000637:Transaminases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D014733:Vibrio / Q000201:enzymology*
D000588:Amines; D004791:Enzyme Inhibitors; D007659:Ketones; D019289:Pyruvic Acid; D007770:L-Lactate Dehydrogenase; D000637:Transaminases; D000409:Alanine