毕赤酵母菌分泌表达重组抑肽酶的高密度发酵工艺研究-东南大学学报医学版

毕赤酵母菌分泌表达重组抑肽酶的高密度发酵工艺研究

单位：中国药科大学

分类号：

出版年·卷·期（页码）：2011·30·第五期（744-748）

摘要：

目的: 对重组毕赤酵母高密度表达抑肽酶的发酵工艺进行探讨。方法: 先通过摇瓶培养方法，确定表达抑肽酶的毕赤酵母在BSM培养基中生长所需几种重要配方的最佳浓度：甘油、氨水、及诱导阶段甲醇的流加量。在此基础上，采用分批补料培养方法，对重组毕赤酵母pSA/GS115 进行高密度发酵。结果: BSM培养基中生长必需的几种重要配方的最佳浓度为：甘油40 g/L、氨水单次补加量0.1%、甲醇流加量1%。经过毕赤酵母高密度发酵，发酵液上清中蛋白的表达量为37.08 mg/L，目的蛋白活性达到4450 BAEEU/ml，比摇瓶培养表达提高了8倍。结论: 成功实现了酵母工程菌的高密度表达发酵，为下一步规模化制备抑肽酶奠定了基础。

Objective: To evaluate the expression of aprotinin in recombinant Pichia pastoris by the high density fermentation. Methods: We identified several important composition of BSM medium of aprotinin Pichia pastoris using shaking flask. The above described optimal conditions were used for scaling up in 7L fermentor. Engineering bacteria pSA/GS115 was fermented in fed-batch high density fermentation. Results: The optimal levels of BSM medium were glycerol 40 g/L, ammonia water 0.1%, methanol 1%. After the high density fermentation, The yield of protein was 37.08 mg/ml. The biological activity determined by ultraviolet absorption assay was 4450BAEEU/ml, which was 8 folds higher compared to shaking flask. Conclusion: All data imply that engineering bacteria was fermented in fed-batch high-density fermentation successfully, which made a foundation for the large- scale production of aprotinin.

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