As a native CoQ10 producer, Rhodobacter sphaeroides has been extensively engineered to enhance CoQ10 production. However, the genetic manipulations using plasmids suffer from risk of plasmid loss during propagation process, biomass impairment due to cellular burden and bio-safety concerns. In this paper, genomic manipulations via Tn7 transposition was conducted to boost the CoQ10 biosynthesis in R. sphaeroides. The titer production and content of CoQ10 were improved by 18.44% and 18.87%, respectively compared to the wild type, when an additional copy of dxs and dxr were integrated into the genome. Further overexpression of idi and ispD by genomic integration created strain RSPCDDII with CoQ10 production and content of 81.23 mg/L and 5.93 mg/g, which were 54.28 and 55.97% higher than those of the wild type. The gene segments were successfully inserted into the attTn7 site of the R. sphaeroides genome. Meanwhile, the biomass was not affected. Compared to overexpression of genes on plasmids, this strategy could enhance protein expression to a proper level without affecting cell growth, and in a more stable manner.
Keywords: Rhodobacter sphaeroides; attTn7 site; CoQ10; genomic integration; metabolic engineering.
© The Author(s) 2022. Published by Oxford University Press on behalf of FEMS.