Abstract

Coenzyme Q10 is required for respiratory electron transport and protects biological membranes against oxidative damage. As coenzyme Q10 supplements are used to treat or to alleviate symptoms associated with an increasing number of health conditions, there is growing interest in the development of bioprocesses for its production. The biosynthesis of coenzyme Q10 involves the condensation of an isoprenoid, decaprenyl diphosphate, with an aromatic compound, para-hydroxybenzoate, followed by a series of modifications of the aromatic moiety of the molecule via the ubiquinone pathway. Escherichia coli naturally produces coenzyme Q8, but replacement of its octaprenyl diphosphate synthase by a decaprenyl diphosphate synthase is sufficient to eliminate the production of coenzyme Q8 and favor the synthesis of coenzyme Q10.
A rational genetic engineering approach was used to create a strain of E. coli capable of producing high levels of coenzyme Q10. First, the endogenous octaprenyl diphosphate synthase gene was deleted and functionally replaced by a decaprenyl diphosphate synthase-encoding gene derived from Sphingomonas baekryungensis. Additionally, this strain was engineered to produce elevated levels of para-hydroxybenzoate by over-expressing genes encoding enzymes of the E. coli shikimate pathway. The production of isoprenoid was increased by introducing a heterologous mevalonate pathway. Decaprenyl diphosphate and para-hydroxybenzoate were further directed toward the ubiquinone pathway by overexpressing a para-hydroxybenzoate prenyltransferase. The resulting recombinant strain was capable of producing elevated levels of coenzyme Q10. In order to further enhance production of this antioxidant, an investigation into the interplay between coenzyme Q10 biosynthesis and primary metabolism was conducted. This investigation revealed a link between sorbitol catabolism and coenzyme Q10 production in the engineered strain. Moreover, abrogating carbon flux to acetate by selected gene knock-outs also enhanced coenzyme Q10 accumulation. However, the engineered strains developed through this research were found to be highly unstable, leading to high variability in coenzyme Q10 production. This instability was hypothesized to result from the burden exerted by the engineered aromatic and foreign mevalonate pathways on primary metabolism. As a result, further optimization of these engineered strains of E. coli will be required in order to develop a suitable platform for coenzyme Q10 production.