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首页> 美国卫生研究院文献>Frontiers in Bioengineering and Biotechnology >Build Your Bioprocess on a Solid Strain—β-Carotene Production in Recombinant Saccharomyces cerevisiae
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Build Your Bioprocess on a Solid Strain—β-Carotene Production in Recombinant Saccharomyces cerevisiae

机译：在固体菌株上构建您的生物过程-重组酿酒酵母中的β-胡萝卜素生产

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Robust fermentation performance of microbial cell factories is critical for successful scaling of a biotechnological process. From shake flask cultivations to industrial-scale bioreactors, consistent strain behavior is fundamental to achieve the production targets. To assert the importance of this feature, we evaluated the impact of the yeast strain design and construction method on process scalability -from shake flasks to bench-scale fed-batch fermentations- using two recombinant Saccharomyces cerevisiae strains capable of producing β-carotene; SM14 and βcar1.2 strains. SM14 strain, obtained previously from adaptive evolution experiments, was capable to accumulate up to 21 mg/gDCW of β-carotene in 72 h shake flask cultures; while the βcar1.2, constructed by overexpression of carotenogenic genes, only accumulated 5.8 mg/gDCW of carotene. Surprisingly, fed-batch cultivation of these strains in 1L bioreactors resulted in opposite performances. βcar1.2 strain reached much higher biomass and β-carotene productivities (1.57 g/L/h and 10.9 mg/L/h, respectively) than SM14 strain (0.48 g/L/h and 3.1 mg/L/h, respectively). Final β-carotene titers were 210 and 750 mg/L after 80 h cultivation for SM14 and βcar1.2 strains, respectively. Our results indicate that these substantial differences in fermentation parameters are mainly a consequence of the exacerbated Crabtree effect of the SM14 strain. We also found that the strategy used to integrate the carotenogenic genes into the chromosomes affected the genetic stability of strains, although the impact was significantly minor. Overall, our results indicate that shake flasks fermentation parameters are poor predictors of the fermentation performance under industrial-like conditions, and that appropriate construction designs and performance tests must be conducted to properly assess the scalability of the strain and the bioprocess.
机译：微生物细胞工厂强大的发酵性能对于成功扩大生物技术过程至关重要。从摇瓶培养到工业规模的生物反应器，一致的应变行为对于实现生产目标至关重要。为了证明这一特征的重要性，我们评估了酵母菌株设计和构建方法对过程可扩展性的影响-从摇瓶到台式规模的分批补料发酵-使用两种能够产生β-胡萝卜素的重组酿酒酵母菌株。 SM14和βcar1.2菌株。先前从适应性进化实验中获得的SM14菌株能够在72 h摇瓶培养物中累积高达21 mg / gDCW的β-胡萝卜素。而通过carotenogenic基因的过表达构建的βcar1.2，仅积累5.8 mg / gDCW的胡萝卜素。令人惊讶的是，这些菌株在1L生物反应器中的分批补料培养导致相反的性能。与SM14菌株（分别为0.48 g / L / h和3.1 mg / L / h）相比，βcar1.2菌株的生物量和β-胡萝卜素生产率（分别为1.57 g / L / h和10.9 mg / L / h）高得多。 。培养80小时后，SM14和βcar1.2菌株的最终β-胡萝卜素效价分别为210和750 mg / L。我们的结果表明，发酵参数的这些实质性差异主要是SM14菌株的Crabtree效应加剧的结果。我们还发现，用于将类胡萝卜素基因整合到染色体中的策略影响了菌株的遗传稳定性，尽管影响很小。总体而言，我们的结果表明，摇瓶发酵参数不能很好地预测工业条件下的发酵性能，因此必须进行适当的结构设计和性能测试，以正确评估菌株和生物工艺的可扩展性。

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