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[Display omitted] • A. faecalis ZS-1 possesses simultaneous bioelectricity generation and CO 2 fixation. • Bioelectricity promotes the formation of electrotrophic bacterial biofilms. • Riboflavin is responsible to inward EET at biocathode during CO 2 -fixing process. • The high expression of ppc gene increases the performance of CO 2 fixation. • Bioelectricity contributes to converting carbon dioxide into lipids in MFC system. The CO 2 fixation mechanism by Alcaligenes faecalis ZS-1 in a biocathode microbial fuel cell (MFC) was investigated. The closed-circuit MFC (CM) exhibited a significantly higher CO 2 fixation rate (10.7%) compared to the open-circuit MFC (OC) (2.0%), indicating that bioelectricity enhances CO 2 capture efficiency. During the inward extracellular electron transfer (EET) process, riboflavin concentration increased in the supernatant while cytochrome levels decreased. Genome sequencing revealed diverse metabolic pathways for CO 2 fixation in strain ZS-1, with potential dominance of rTCA and C4 pathways under electrotrophic conditions as evidenced by significant upregulation of the ppc gene. Differential metabolite analysis using LC-MS demonstrated that CM promoted upregulation of various lipid metabolites. These findings collectively highlight that ZS-1 simultaneously generated electricity and fixed CO 2 and that the ppc associated with bioelectricity played a critical role in CO 2 capture. In conclusion, bioelectricity resulted in a significant enhancement in the efficiency of CO 2 fixation and lipid production. [ABSTRACT FROM AUTHOR]