13TSB_SynBio- Novel Bacterial Hosts for Biobutanol Production

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Energy insecurity, global warming and fluctuations in oil prices and has resulted a rapid increase in the demand for the biofuel, bioethanol. However, the alcohol biobutanol is widely acknowledged as a 'superior biofuel'. It is traditionally produced via a sugar-based fermentation process using a bacterium Clostridium acetobutylicum. The process, however, remains uneconomic due to feedstock prices, which represents 60-80% of production costs.Another member of the same family of bacteria, Clostridium pasteurianum is a robust, fast growing bacterium which is also able to grow on glycerol as well as sugars and produce valuable 3-carbon (1,3 propanediol) and 4-carbon chemicals (butyrate & butanol). The microbe has tremendous potential as a fermentation host but has not been exploited due to its limited substrate range and mixed array of chemical products that result in low butanol yield compared to current commercial. In this project we aim to use synthetic biology to introduce new metabolic pathways for starch utilisation and butanol production into C. pasteurianum with the aim to produce butanol in high yield from starch. Genetic manipulation based on gene knockout will also be used to inactivate competing pathways. The partners have extensive experience with solventogenic Clostridia. Nottingham's CRG has developed an impressive range of gene tools and recently completed the determination of the complete genetic blueprint (genome sequence) of C. pasteuranium. GBL has identified novel pathways for both starch degradation and butanol production from its unique collection of commercial strains and leads recommercialisation efforts for the butanol fermentation. This collaboration between GBL and GRG enables, for the first time, a targeted and rational approach for strain improvement in a novel host using synthetic biology and metabolic engineering. This project clearly demonstrates the potential application of synthetic biology for industrial biotechnology. Fermentation performance from the engineered C. pasteurianum strains will be benchmarked against GBL's best performing commercial strains and if promising taken forward for further development. The project output will also support additional projects with this microbe focused on extending the substrate range to include cellulosic feedstocks and developing glycerol metabolism for the production of 3-carbon chemicals.