Bacterial pathogens such as Salmonella enterica serovar Typhimurium (S. Typhimurium) have to cope with fluctuating oxygen levels during infection within host gastrointestinal tracts. The global transcription factor FNR (fumarate nitrate reduction) plays a vital role in the adaptation of enteric bacteria to the low oxygen environment. Nevertheless, a comprehensive profile of the FNR regulon on the proteome level is still lacking in S. Typhimurium. Herein, we quantitatively profiled S. Typhimurium proteome of an fnr-deletion mutant during anaerobiosis in comparison to its parental strain. Notably, we found that FNR represses the expression of virulence genes of Salmonella pathogenicity island 1 (SPI-1) and negatively regulates propanediol utilization by directly binding to the promoter region of the pdu operon. Importantly, we provided evidence that S. Typhimurium lacking fnr exhibited increased antibiotics susceptibility and membrane permeability as well. Furthermore, genetic deletion of fnr leads to decreased bacterial survival in a Caenorhabditis elegans infection model, highlighting an important role of this regulator in mediating host-pathogen interactions.