Quantitative metabolomics analysis of amino acid metabolism in recombinant Pichia pastoris under different oxygen availability conditions
Quantitative metabolomics analysis of amino acid metabolism in recombinant Pichia pastoris under different oxygen availability conditions
复制标题
DOI：
10.1186/1475-2859-11-83
复制
发表时间：
2012-06-15
影响因子：
6.4
通讯作者：
Ferrer, Pau
中科院分区：
文献类型：
--
作者：
Carnicer, Marc;ten Pierick, Angela;Ferrer, Pau
背景：一些转录研究证明，环境和内在应激因素可以导致酵母生理的全局改变。缺氧已被证明对生长在葡萄糖上的毕赤酵母重组蛋白的表达有有益的影响。此外，转录谱分析显示，氧可用性强烈影响麦角甾醇的生物合成、中心碳代谢和应激反应，特别是未折叠蛋白的反应。为了更好地了解氧可用性和外源蛋白分泌对中心代谢的影响和相互作用，首次对不同氧可用性条件下生长的重组巴斯德酵母菌株的游离氨基酸库进行了定量代谢组学分析。结果：得到的数值表明，细胞内氨基酸库因不同的氧可用性条件而发生显著变化，在氧气限制下，它们的大小整体增加。值得注意的是，尽管外源蛋白产量相对较低（约40-80 μ g Fab/g(DCW).h），但重组蛋白的产量对细胞内氨基酸库的影响有限但显著，与参考菌株相比，生产菌株的氨基酸库普遍减少。然而，观察到的单个氨基酸库的变化与重组蛋白序列中相应的相对丰度无关，而是与整个细胞蛋白质氨基酸组成的变化有关。结论：总的来说，所获得的结果，结合之前的转录组学和蛋白质组学分析，提供了一个系统的代谢指纹，表明氧可用性对重组酵母蛋白生产的影响。
Background: Environmental and intrinsic stress factors can result in the global alteration of yeast physiology, as evidenced by several transcriptional studies. Hypoxia has been shown to have a beneficial effect on the expression of recombinant proteins in Pichia pastoris growing on glucose. Furthermore, transcriptional profiling analyses revealed that oxygen availability was strongly affecting ergosterol biosynthesis, central carbon metabolism and stress responses, in particular the unfolded protein response. To contribute to the better understanding of the effect and interplay of oxygen availability and foreign protein secretion on central metabolism, a first quantitative metabolomic analysis of free amino acids pools in a recombinant P. pastoris strain growing under different oxygen availability conditions has been performed.Results: The values obtained indicate significant variations in the intracellular amino acid pools due to different oxygen availability conditions, showing an overall increase of their size under oxygen limitation. Notably, even while foreign protein productivities were relatively low (about 40-80 mu g Fab/g(DCW).h), recombinant protein production was found to have a limited but significant impact on the intracellular amino acid pools, which were generally decreased in the producing strain compared with the reference strain. However, observed changes in individual amino acids pools were not correlated with their corresponding relative abundance in the recombinant protein sequence, but to the overall cell protein amino acid compositional variations.Conclusions: Overall, the results obtained, combined with previous transcriptomic and proteomic analyses provide a systematic metabolic fingerprint of the oxygen availability impact on recombinant protein production in P. pastoris.