Plasmid DNA Production in Proteome-Reduced

The design of optimal cell factories requires engineering resource allocation for maximizing product synthesis. A recently developed method to maximize the saving in cell resources released 0.5% of the proteome of by deleting only three transcription factors. We assessed the capacity for plasmid DNA (pDNA) production in the proteome-reduced strain in a mineral medium, lysogeny, and terrific broths. In all three cases, the pDNA yield from biomass was between 33 and 53% higher in the proteome-reduced than in its wild type strain. When cultured in fed-batch mode in shake-flask, the proteome-reduced strain produced 74.8 mg L pDNA, which was four times greater than its wild-type strain. Nevertheless, the pDNA supercoiled fraction was less than 60% in all cases. Deletion of increased the pDNA yields in the wild type, but not in the proteome-reduced strain. Furthermore, mutants produced a higher fraction of supercoiled pDNA, compared to their parents. These results show that the novel proteome reduction approach is a promising starting point for the design of improved pDNA production hosts.