Decoupling acidogenic cascade via bio-electrogenic induced ABE pathway for enhanced n-butanol synthesis in Clostridium acetobutylicum ATCC-824: Bioenergetics and gene expression analysis

This investigation scrutinized the quantitative aspects of biobutanol production in conventional and bioelectrogenic induced ABE fermentation using Clostridium acetobutylicum ATCC-824. The primary objective of the study was to investigate how alterations in the extracellular redox potential (-0.4 V vs. Ag/AgCl (3.5 M KCl)) affect the hetero-fermentation towards the production of acetone, butanol, and ethanol during the bi-phasic fermentation, utilizing different glucose concentrations from 20 g/L - 60 g/L. The study showed significant results with 68.86 % increased butanol titer (6.67 g/L) compared to conventional fermentation (3.95 g/L butanol). Additionally, the acetone production also increased to 2.34 g/L as compared to conventional fermentation (1.96 g/L). Furthermore, gene expression and network analysis showed that various ABE pathway genes were significantly up or down regulated during the bi-phasic fermentation, indicating a difference between the conventional and EF reactors. The external potential influenced the overexpression of genes like pta, crt, bcd, ask, adh (CA_C1574), and hbd (CA_C3375), while simultaneously downregulation of buk gene, suggesting the acidogenic decoupling and transition from cold channel (butyric acid) towards hot channel pathway (directly from acetyl CoA through butyryl-CoA) for improved butanol production. Moreover, the study also revealed that entropy and enthalpy changes played a significant role in butanol production and ABE fermentation. The EF showed more negative free energy (-3.33 x 104 kJ/mol) when compared with conventional fermentation (-2.8 x 104 kJ/mol) suggesting the mechanism to be more spontaneous and viable. Overall, this study provided evidence of metabolic rerouting with non-genetic approach by regulating electron flow, culminating in an impact on the bi-phasic fermentation towards specific metabolites.