Genetic engineering and strain optimization of Clostridium ljungdahlii for the production of biobutanol by syngas fermentation

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Worldwide the energy demands are increasing and sources of fossil energy carriers are limited. Routes to produce renewable energy, transportation fuels and commodities have to be developed. The most prominent biofuel used today is ethanol produced from sugar cane, corn or wheat (1st generation fuel). Instead of using these food plants for the production of biofuels, lignocellulosic biomass is considered to play an important role in overcoming problems associated with 1st generation fuels such as land use conflicts. Furthermore, n-butanol as an alternative to ethanol offers advantages such as a higher energy density and better combustion properties. There are two platforms described for the production of 2nd generation biofuels from lignocellulose: the sugar and the synthesis gas (syngas) platform. In the sugar platform, lignocellulose is converted into sugars and lignin by extensive pretreatment with heat, chemicals, and enzymes, and the sugars are further fermented into bioalcohols. The syngas platform (thermochemical route) uses gasification or slow pyrolysis of the indigestive lignocellulosic biomass to produce syngas, a mixture of mainly carbon monoxide (CO), hydrogen (H2), carbon dioxide (CO2) and nitrogen (N2). The produced syngas can then be converted into medium to long chain alcohols in the thermo-chemical Fischer-Tropsch process. While in this process chemical catalysts are poisoned by impurities in the gas and the products are depending on the gas mixture composition, alternatively, carboxydotrophic bacteria can be used to specifically produce multicarbon compounds, such as acetate, ethanol, or n-butanol, in a process called syngas fermentation.Considerable progress is made in reactor design to use carboxidotrophic, homoacetogenic bacteria for the fermentation of syngas into bioalcohols and/ or acetate using the syngas platform. The production of n-butanol is still challenging. Another big challenge is the lack of sufficient genetic tools to optimize these organisms via genetic engineering.The proposed work aims to establish syngas fermentation to produce n-butanol as a 2nd generation biofuel and/ or platform chemical and optimize the process through: 1) optimization of the syngas fermenting organism Clostridium ljungdahlii by genetic engineering; and 2) the optimization of fermentation conditions such as co-feeding of syngas and the carboxylate substrate n-butyrate.