Biooxidation of n-octane by a recombinant Escherichia coli in a two-liquid-phase system: Effect of medium components on cell growth and alkane oxidation activity

Abstract

When Escherichia coli hosts are transformed with the alk genes, which encode the alkane oxidation pathway of Pseudomonas oleovorans, the recombinants are capable of converting alkanes to alkanoic acids during growth in two-liquid-phase (aqueous-organic) media. In such media, the cells grow on carbohydrates which are dissolved in the water phase, while the alkane oxidation system converts the alkanes of the nonaqueous phase to alkanoates which solubilize mainly in the aqueous phase. The performance of this system depends on the nature of the aqueous carbon source, the presence of growth-promoting additions in the aqueous phase, the presence of a bulk apolar alkane phase, the presence in the aqueous phase of various acids produced during the fermentation, and the state of the cells. These effects have been explored using E. coli W3110, into which the plasmid pGEc47, which encodes the alk genes from the catabolic OCT plasmid from P. oleovorans, has been introduced. The aqueous carbon source used played a key role in the efficiency of the biooxidation of n-alkanes. When E. coli was cultivated in medium with yeast extract, the specific bioconversion of n-octane to n-octanoate reached 65 μmol min −1 g −1 of cells, whereas in the medium without addition of yeast extract the activity remained at 15 μmol min −1 g −1 of cells. Induction of the expression of the alk genes in the recombinant E. coli altered cell physiology, which resulted in a limitation of the final cell dry weight. Under optimized conditions, a bioreactor productivity of 3.6 mmol h −1 l −1 could be maintained for 12 h resulting in a final concentration of 25 m m n-octanoate in the aqueous phase.