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首页> 外文期刊>Journal of Biotechnology >Monitoring of transcriptome and proteome profiles to investigate the cellular response of E. coli towards recombinant protein expression under defined chemostat conditions
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Monitoring of transcriptome and proteome profiles to investigate the cellular response of E. coli towards recombinant protein expression under defined chemostat conditions

机译：监测转录组和蛋白质组图谱，以研究大肠杆菌在规定的化学恒温条件下对重组蛋白表达的细胞反应

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The use of strong promoter systems for recombinant protein production generates high product yields, but also overburdens the host cell metabolism and compromises production. Escherichia coli has highly developed regulatory pathways that are immediately responsive to adverse conditions. To gain insight into stress response mechanisms and to detect marker genes and proteins for stress specific monitoring time course analysis of controlled chemostat cultivations was performed using E. coli total microarray and difference gel electrophoresis (Ettan(TM) DIGE). In order to detect differences and consistencies of stress response as well as the impact of the inducer isopropyl-beta-d-thiogalactopyranosid on cells, expression of two recombinant proteins (hSOD and GFPmut3.1) was investigated. Genes involved in aerobic metabolism under control of the ArcB/ArcA two component system were found to be down-regulated, and the interplay of the psp operon, ArcA system and guanosine tetraphosphate is suggested to be involved in stress regulatory mechanisms. A distinct impact of the two recombinant proteins was observed, particularly on levels of known stress regulatory genes and proteins, as well as on the response associated with ArcA and psp. Altogether, 62 genes as well as seven proteins showed consistent expression levels due to recombinant gene expression, and are therefore suggested to be appropriate monitoring targets.
机译：在重组蛋白生产中使用强启动子系统可产生高产量，但同时也使宿主细胞的代谢负担过重，并损害了生产。大肠杆菌具有高度发达的调节途径，可立即对不利条件作出反应。为了深入了解应激反应机制并检测标记基因和蛋白质以进行应激特异性监测，使用大肠杆菌总微阵列和差异凝胶电泳（Ettan TM DIGE）对受控的恒化器培养进行了时程分析。为了检测应激反应的差异和一致性，以及诱导剂异丙基-β-d-硫代吡喃半乳糖苷对细胞的影响，研究了两种重组蛋白（hSOD和GFPmut3.1）的表达。发现在ArcB / ArcA两组分系统的控制下参与有氧代谢的基因被下调，而psp操纵子，ArcA系统和鸟苷四磷酸的相互作用被认为参与了压力调节机制。观察到两种重组蛋白的显着影响，特别是对已知的应激调节基因和蛋白的水平以及与ArcA和psp相关的反应的影响。由于重组基因的表达，总共有62个基因以及7种蛋白质显示出一致的表达水平，因此建议作为合适的监测目标。

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