RESUMO
Production of chiral amines and unnatural amino-acid using ω-transaminase can be achieved by kinetic resolution and asymmetric synthesis, thus ω-transaminase is of great importance in the synthesis of pharmaceutical intermediates. By genomic data mining, a putative ω-transaminase gene hbp was found in Burkholderia phytofirmans PsJN. The gene was cloned and over-expressed in Escherichia coli BL21 (DE3). The recombinant enzyme (HBP) was purified by Ni-NTA column and its catalytic properties and substrate profile were studied. HBP showed high relative activity (33.80 U/mg) and enantioselectivity toward ß-phenylalanine (ß-Phe). The optimal reaction temperature and pH were 40 â and 8.0-8.5, respectively. We also established a simpler and more effective method to detect the deamination reaction of ß-Phe by UV absorption method using microplate reader, and demonstrated the thermodynamic property of this reaction. The substrate profiling showed that HBP was specific to ß-Phe and its derivatives as the amino donor. HBP catalyzed the resolution of rac-ß-Phe and its derivatives, the products (R)-amino acids were obtained with about 50% conversions and 99% ee.