表达胰岛素的毕赤酵母生长动力学及诱导策略

Studies of the growth kinetics and induction strategy with a strain of recombinant yeast Pichia Pastoris expressing the human insulin gene

对本实验室构建的一株重组巴斯德毕赤酵母基因工程菌的生长及诱导进行了研究。结果表明,生长阶段菌体的生长与限制性基质甘油残留浓度的关系符合Monod关系式,细胞最大生长比速率为0.204 h-1,饱和常数为24.3 g/L。经参数推导得理论最大细胞对甘油得率为0.497 g/g,以及菌体生长维持系数为0.007 g/g.h,后者说明此株工程菌有高密度发酵的潜力。诱导阶段补加酵母提取物、大豆蛋白胨和微量元素能有效提高目的蛋白产量。在发酵罐上罐试验中,目的蛋白产量达256 mg/L,蛋白产量是摇瓶发酵蛋白产量的5倍多。

The growth kinetics and induction strategy of a genetically engineered yeast Pichia Pastoris which expresses the human insulin gene have been studied. The results indicate that the relationship between the specific growth rate （μ） and the residual concentration of glycerol is in accord with the Monod equation in the cell growth phase where μmax = 0. 204 h^-1, and Ks = 24.3 g/L. The maximum cell yield of glycerol is 0. 497 g/g, and the growth maintaining coefficient rn is 0. 007 g/g· h. The latter value indicates that the strain has the potential for high density cultivation. It was found that the heterologous protein concentration could be increased by using a batch-fed yeast extract, with soyabean peptone and trace elements added in the induction phase. The heterologous protein concentration reached 256 mg/L in fermentation in a 10 L fermenter, which is 5 times the level obtained in flask fermentation. The results may be a guide to future industrial production.