重组巴斯德毕赤酵母发酵生产S-腺苷蛋氨酸

S-adenosyl-L-methionine Production by Recombinant Pichia pastoris

目的:考察L-甲硫氨酸、甲醇、pH值、接种量、装液量、油酸以及PTM1 7个因子对重组巴斯德毕赤酵母发酵生产S-腺苷蛋氨酸的影响。方法:首先通过部分因子试验设计,确定影响S-腺苷蛋氨酸产量的关键因子,然后通过Box-Behnken试验设计,确定其生产S-腺苷蛋氨酸的最佳条件。结果:影响重组巴斯德毕赤酵母发酵生产S-腺苷蛋氨酸的关键因子为甲醇、pH值和装液量。当甲醇添加量1.5%,pH值5.0,装液量30 mL/500 mL时,S-腺苷蛋氨酸的产量达到最高值。优化后的最佳培养基组成为:1.00%酵母膏,2.00%蛋白胨,1.34%YNB,1.0%L-met,100 mmol/L磷酸盐缓冲液,1.40%甲醇,0.10%油酸,0.4%PTM1以及4.00×10^(-5)%生物素。在优化条件下培养84 h后,S-腺苷蛋氨酸的产量达到最高值3.23 mg/mL,是优化前的1.8倍。结论:本研究结果为S-腺苷蛋氨酸的产业化生产提供了良好基础。

Objective：To explore the effect of seven factors,including L-met,methanol,pH,inoculation,fermentation broth volume,oleic acid and PTM1,on the S-adenosyl-L-methionine production by recombinant P.pastoris.Methods：Fractional factorial design was used to determine the key factors that affected the production of S-adenosyl-L-methionine,and then the optimal culture conditions were determined by Box-Behnken design.Results：Methanol,pH and fermentation broth volume were found to be the key factors,and their optimal values were 1.5%,5.0 and 30 mL/500 mL,respectively.The optimal medium components were as follows ：yeast extract 1.00%,peptone 2.00%,YNB1.34%,L-met 1.0%,phosphate buffer 100 m M,methanol 1.4%,oleic acid 0.10%,PTM10.4% and biotin 4.00×10^-5%.Under these conditions,the highest yield of S-adenosyl-L-methionine was up to 3.23 mg/mL after P.pastoris was cultured for 84 h,which was 1.8-fold higher than that obtained under the primitive conditions.Conclusion：This study lays a good foundation for future industrial production of S-adenosyl-L-methionine.