Genetic Manipulation of a Twin Clostridia Consortium for Co-production of n-Butanol and Isobutanol by Consolidated Bioprocessing

It is attractive to co-produce n-butanol and isobutanol from lignocellulosic biomass in engineered strains or consortia, because they can be coseparated and combined for use as biofuel. However, very few attempts have been made to date. To address that, a heterologous isobutanol pathway was first introduced to enable Clostridium beijerinckii to coproduce n-butanol and isobutanol. Following medium optimization, the recombinant C. beijerinckii produced 194 mg/L isobutanol and 7.16 g/L n-butanol from glucose. C. cellulovorans was then genetically engineered by overexpressing adhE1, kivD, and yqhD to synthesize n-butanol and isobutanol with alkali-extracted, deshelled corn cobs (AECC) as the sole carbon source. It produced 156 mg/L isobutanol and 1.81 g/L n-butanol from 39.5 g/L AECC within 120 h. Finally, the above two recombinant strains were mix-cultured and optimized. The optimal consortium degraded 55.1 g/L AECC and produced 1.05 g/L isobutanol and 6.22 g/L n-butanol, which were 6.73-and 3.44-fold higher than those obtained from single culture of recombinant C. cellulovorans. Besides, the titer and yield of total butanol are 15.6% and 11.9%, respectively, higher than those of recombinant consortia that only produce n-butanol. Synthetic biology empowers artificial consortia with expanded functions, as well as improved performance, showing great potential in consolidated bioprocessing.