| 摘要： |
Soybean seed coat peroxidase (SBP) is a valuable enzyme having a broad variety of applications in analytical chemistry, biochemistry, and food processing. In the present study, the sscp gene (Gene ID: 548068) was optimized based on the preferred codon usage of Escherichia coli, synthesized, and expressed in E. coli BL21(DE3). SDS-PAGE and western blot analysis of this expressed protein revealed that its molecular weight is approximately 39kDa. The effects of induction temperature, concentration of isopropyl--D-thiogalactoside and hemin, induction time, expression time were optimized to enhance SBP production with a maximum activity of 11.23U/mL (8.64U/mg total protein). Furthermore, the kinetics of enzyme-catalyzed reactions of recombinant protein was determined. When 2,2-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) was used as substrate, optimum reaction temperature and pH of the enzyme were 85 degrees C and 5.0, respectively. The effects of metal ions on the enzymatic reaction were also further investigated. The SBP was successfully expressed in E. coli BL21(DE3) which would provide a more efficient production strategy for industrial applications of SBP. |
