Abstract: In order to synthesize high value-added β-phenylalanine, the gene encoding phenylalanine aminomutase from Taxus chinensis was cloned and expressed in Escherichia coli. The expression vector Pet-sumo-Tcpam was successfully constructed and transferred into E. coli BL21 to express the recombinant enzyme (TcPAM). Electrophoretically pure TcPAM was obtained using affinity chromatography. The results of mass spectrometry (MS) and circular dichroism (CD) spectroscopy showed that TcPAM could catalyze isomerization of α-phenylalanine to R-β-phenylalanine. Its activity was 4.11 U/mg under the optimum conditions of 30 ℃ and pH 9. The metal ions K+, Fe2+ and Ca2+ as well as the surfactants cetrimonium bromide (CTAB), sodium dodecyl sulphate (SDS), Triton X-100, and Tween 80 had little effect on its activity, resulting in retention of about 90% of the initial activity, while Cu2+ and Zn2+ had strong inhibitory effect on TcPAM, Furthermore, when TcPAM was used to catalyze isomerization of α-arylalanines with different groups on the benzene ring to R-β-phenylalanine, the presence of electron-donating groups on the benzene ring promoted transfer of the α-amino group to the β site as compared to the presence of electron-withdrawing groups, improving the substrate conversion rate and the highest value of 45% was obtained when the R group was 4-MeO or 4-Me. This study provides the basis for enzymatic synthesis of β-arylalanine.

Key words: Taxus chinensis; phenylalanine ainomutase; β-arylalanine