Triterpene hydrocarbon production engineered into a metabolically versatile host-Rhodobacter capsulatus

Triterpene hydrocarbon biosynthesis of the ancient algae Botryococcus braunii was installed into Rhodobacter capsulatus to explore the production of C₃₀ hydrocarbon in a host capable of diverse growth habits-utilizing carbohydrate, sunlight or hydrogen (with CO₂ fixation) as alternative energy feedstocks. Engineering an enhanced MEP pathway was also used to augment triterpene accumulation. Despite dramatically different sources of carbon and reducing power, nearly the same level of botryococcene or squalene (∼5mg oil/g-dry-weight [gDW]) was achieved in small-scale aerobic heterotrophic, anaerobic photoheterotrophic, and aerobic chemoautotrophic growth conditions. A glucose fed-batch bioreactor reached 40mg botryococcene/L (∼12mg/gDW), while autotrophic bioreactor performance with CO₂, H₂, and O₂ reached 110mg/L (16.7mg/gDW) during batch and 60mg/L (23mg/gDW) during continuous operation at a dilution rate corresponding to about 10% of μ_max. Batch and continuous autotrophic specific productivity was found to reach 0.5 and 0.32mg triterpene/g DW/h, comparable to prior reports for terpene production driven by heterotrophic growth conditions. This demonstrates the feasibility of alternative feedstocks and trophic modes to provide comparable routes to biochemicals that do not rely on sugar.