A comparative study of global stress gene regulation in response to overexpression of recombinant proteins in Escherichia coli.
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% (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), SOS/DNA damage (recA, lon, IS5 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.
Gill
R T
RT
Department of Chemical Engineering, University of Maryland, College Park, Maryland 20742, USA.
Valdes
J J
JJ
Bentley
W E
WE
D010582:Bacteriophage lambda / Q000235:genetics; D001698:Biomedical Engineering; D004249:DNA Damage; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D018869:Heat-Shock Response / Q000235:genetics; D010641:Phenotype; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013014:SOS Response, Genetics / Q000235:genetics
D011994:Recombinant Proteins