(571f) Metabolic Engineering of Clostridium Tyrobutyricum for Isopropanol Production

Isopropanol is one of the most widely used solvents in the world. It is also one of the secondary alcohols that can be used as a direct or partial replacement for gasoline. Isopropanol can be produced by anaerobic microorganisms such as Clostridium beijerinckii. However, the native metabolic pathways are limited by tight regulation and the isopropanol production is low, less than 30 mM, in these strains. Clostridium tyrobutyricum is an anaerobic acidogen which can convert glucose and xylose to acetate and butyrate. Previously, our group had engineered this bacterium to produce n-butanol at a high rate, yield, and titer that are even higher than most of those reported native solventogenic Clostridia. In this work, we studied the feasibility of enabling C. tyrobutyricum to produce isopropanol. A plasmid was constructed with C. acetobutylicum ctf (acetoacetyl-CoA transferase), adc (acetoacetate decarboxylase) and C. beijerinckii adh (secondary alcohol dehydrogenase) genes under C. tyrobutyricum ATCC 25755 thl (acetoacetyl-CoA transferase) promoter. The plasmid was transformed into both C. tyrobutyricum wild type and ack (acetate kinase) knockout mutant strains. Isopropanol production by these mutants were confirmed in batch fermentation with glucose as the carbon source. The effects of isopropanol production on cell growth and fermentation kinetics were studied and will be presented in this paper.