A Comparative Study of Global Stress Gene Regulation in Response to Overexpression of Recombinant Proteins in Escherichia coli

✍ Scribed by R.T. Gill; J.J. Valdes; W.E. Bentley

• Publisher: Elsevier Science
• Year: 2000
• Tongue: English
• Weight: 353 KB
• Volume: 2
• Category: Article
• ISSN: 1096-7176
• DOI: 10.1006/mben.2000.0148

✦ Synopsis

Global gene regulation throughout the Escherichia coli stress response to overexpression of each of five recombinant proteins was evaluated. Reverse-transcriptase polymerase chain reaction-amplified mRNA from induced and control cells were hybridized with a DNA array of Kohara clones representing 16 0 (700 genes) of the E. coli genome. Subsequently, Northern analysis was performed for quantification of specific gene dynamics and statistically significant overlap in the regulation of 11 stress-related genes was found using correlation analysis. The results reported here establish that there are dramatic changes in the transcription rates of a broad range of stress genes (representing multiple regulons) after induction of recombinant protein. Specifically, the responses included significantly increased upregulation of heat shock ( ftsH , clpP, lon, ompT, degP, groEL, aceA, ibpA), SOSÂ DNA damage (recA, lon, IS< transposase), stationary phase (rpoS, aceA), and bacteriophage life cycle ( ftsH , recA) genes. Importantly, similarities at the microscopic (gene) level were not clearly reflected at the macroscopic (growth rate, lysis) level. The use of such dynamic data is critical to the design of gene-based sensors, the engineering of metabolic pathways, and the determination of parameters (harvest and induction times) needed for successful recombinant E. coli fermentations.