D-阿洛酮糖 3-差向异构酶基因在枯草芽孢杆菌中的表达

Expression of D-Psicose 3-Epimerase in Bacillus subtilis

摘要:

将D-阿洛酮糖3-差向异构酶(DPE)基因利用PCR进行扩增, 与枯草芽孢杆菌载体pMA5连接, 构建重组质粒pMA5-cbdpe。重组质粒转入枯草芽孢杆菌Bacillus subtilis WB800感受态细胞, 利用卡那霉素筛选和PCR鉴定, 获得一株DPE重组枯草芽孢杆菌菌株。该重组菌株无需诱导即可产生DPE酶, 18 h时酶活可达6.8 U/mL。该酶最适pH为7.0, 最适温度为55 ℃, 与大肠杆菌表达的DPE酶酶学性质相似。结果表明, DPE酶可在枯草芽孢杆菌中表达。

Abstract:

The D-psicose 3-epimerase gene was amplified through PCR and ligated with the expression vector pMA5 of Bacillus subtilis, forming the recombinant plasmid pMA5-cbdpe. The recombinant plasmid was then transformed into B. subtilis WB800, and a recombinant strain was acquired through resistance screening of Kanamycin and PCR identification method. The enzyme activity of recombinant D-psicose 3-epimerase could reach 6.8 U/mL after 18 h fermentation. The recombinant D-psicose 3-epimerase expressed in B. subtilis exhibited optimal activity at pH 7.0. And the optimal temperature was 55 ℃. It showed the same enzyme characterization as the D-psicose 3-epimerase expressed in Escherichia coli. This result indicated that the D-psicose 3-epimerase gene was successfully expressed in B. subtilis.