Journal
ENZYME AND MICROBIAL TECHNOLOGY
Volume 39, Issue 3, Pages 391-398Publisher
ELSEVIER SCIENCE INC
DOI: 10.1016/j.enzmictec.2005.11.048
Keywords
metabolic burden; DH5 alpha; fermentation; plasmid maintenance
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DNA plasmids of Escherichia coli are common vectors for recombinant protein and metabolite production and have potential therapeutic applications as genetic vaccines and therapeutics. However, plasmid maintenance imposes a metabolic burden on the host cells, resulting in reduced growth rate and cell density. In 2 L batch fermentation, DH5 alpha cells carrying a 7.3 kb NS3 plasmid had a lower specific growth rate than the non-plasmid-bearing host (0.64 h(-1) versus 0.87 h(-1)). In this work, global transcriptional analysis was combined with proteomics studies to evaluate the effect of plasmid maintenance on gene expression. Global transcriptional expression analysis of plasmid-bearing cells over host showed a general trend of downregulated biosynthetic/energy metabolism genes, differentially expressed transport genes and upregulated heat shock proteins. In the central metabolic pathways, most glycolytic genes were downregulated, while less expression difference was found in the pentose phosphate pathway. Expression ratios of 19 proteins identified from proteomics studies were consistent with these observations. Our findings suggest that plasmid maintenance alone perturbs global gene regulation, and leads to significant changes in central metabolic pathways in the host. This work contributes to our understanding of plasmid metabolic load at the gene expression level and could potentially aid in future metabolic engineering efforts. (c) 2006 Elsevier Inc. All rights reserved.
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