响应面法优化甲硫氨酸γ-裂解酶发酵培养基

甘祥武; 胡海艳; 黄魁英; 陈舒洁; 李学优

响应面法优化甲硫氨酸γ-裂解酶发酵培养基

Optimization of fermentation medium of methionine γ-lyase by using response surface methodology

摘要

摘要: 目的:为获得低成本高酶活的工业化培养基,对甲硫氨酸γ-裂解酶(MGL)发酵生产培养基进行优化。方法:通过碳、氮源的筛选确定对酶活具有明显影响的因子,通过Plackett-Burman设计确定主要影响因子:甘油、安琪酵母粉、智龙酵母粉;采用最陡爬坡实验及Box-Behnken响应面设计确定了三个主要影响因子的最适条件,当甘油浓度为1.40%,安琪酵母粉为0.80%,智龙酵母粉为0.96%时,甲硫氨酸裂解酶酶活达到最高值为11.30 U/mL。结果:经过发酵验证,MGL酶活为(11.06±0.15) U/mL,较LB培养基1.65 U/mL提高了5.69倍,显示出了良好的优化效果。结论:经优化后最终培养基组成为:甘油1.40%、安琪酵母粉0.80%、智龙酵母粉0.96%、乳糖1.20%、安琪蛋白胨1.80%、NaCl 0.90%、K₂HPO₄ 1.80%、MgSO₄ 0.20%。

Abstract: Objective:In order to obtain low-cost and high-activity industrial medium,the fermentation medium of methionine γ-lyase(MGL)was optimized. Method:The factors influencing the enzyme activity were determined by single factor experiment,The factors influencing the activity of the enzyme were investigated by Plackett-Burman design.The result showed that the concentration of glycerol,Angel yeast,zhilong yeast played important roles in influencing the content of MGL.The most significant conditions were determined by the steepest climbing experiment and Box-Behnken response surface design. Result:The results showed that the activity of MGL reached 11.30 U/mL when the concentration of glycerol was 1.40%,Angel yeast was 0.80% and zhilong yeast was 0.96%. After fermentation verification,the activity of MGL was(11.06±0.15) U/mL,which was 5.69 times higher than LB medium 1.65 U/mL,showing a good optimization effect. Conclusion:The optimal proliferation medium was as follows:glycerol 1.40%,Angel yeast 0.80%,zhilong yeast 0.96%,lactose 1.20%,Angel peptone 1.80%,NaCl 0.90%,K₂HPO₄ 1.80%,MgSO₄ 0.20%.