Aerobic acetone-butanol-isopropanol (ABI) fermentation through a co-culture of Clostridium beijerinckii G117 and recombinant Bacillus subtilis 1A1

An engineered B. subtilis 1A1 strain (BsADH2) expressing a secondary alcohol dehydrogenase (CpSADH) was co-cultured with C. beijerinckii G117 under an aerobic condition. During the fermentation on glucose, B. subtilis BsADH2 depleted oxygen in culture media completely and created an anaerobic environment for C. beijerinckii G117, an obligate anaerobe, to grow. Meanwhile, lactate produced by B. subtilis BsADH2 was re-assimilated by C. beijerinckii G117. In return, acetone produced by C. beijerinckii G117 was reduced into isopropanol by B. subtilis BsADH2 via expressing the CpSADH, which helped maintain the redox balance of the engineered B. subtilis. In the symbiotic system consisting of two strains, 1.7 g/L of acetone, 4.8 g/L of butanol, and 0.9 g/L of isopropanol (with an isopropanol/acetone ratio of 0.53) was produced from 60 g/L of glucose. This symbiotic system also worked when oxygen was supplied to the culture, although less isopropanol was produced (0.9 g/L of acetone, 4.9 g/L of butanol, and 0.2 g/L of isopropanol). The isopropanol titer was increased substantially to 2.5 g/L when we increased the inoculum size of B. subtilis BsADH2 and optimized other process parameters. With the Bacillus-Clostridium co-culture, switching from the original acetone-butanol (AB) fermentation to an aerobic acetone-butanol-isopropanol (ABI) fermentation can be easily achieved without genetic engineering of Clostridium. This strategy of employing a recombinant Bacillus to co-culture with Clostridium should be potentially useful to modify traditional acetone-butanol-ethanol fermentation for the production of other value-added chemicals.