Regulation of DbaM modifies the disulfide bond formation system in Escherichia coli.

Escherichia coli YmdA protein modulates biofilm formation and susceptibility to the aminoglycoside antibiotic apramycin (APR). Here, we renamed this protein DbaM (Dsb-associated Biofilm and Antibiotic Susceptibility Modulatory protein) to reflect its physiological role. We show that DbaM function requires translocation to the periplasm via its N-terminal signal sequence and subsequent Dsb-dependent maturation. The formation of an intramolecular disulfide bond between conserved cysteine residues (C41 and C54) is required for activity; mutations at these sites negate biofilm inhibition and APR sensitization. Rather than forming a stable complex, DbaM modulates the catalytic turnover of the disulfide bond (Dsb) network, likely through transient and catalytic mixed disulfide intermediates. Interestingly, we demonstrated that periplasmic redox modulation by disulfide bond formation protein A (DsbA)-associated action is dependent on DbaM activity. This process involves both the DsbA oxidative and disulfide bond isomerase C (DsbC) pathways and alters the periplasmic thiol-disulfide balance toward net oxidation. Additionally, oxidized DbaM structurally mimics major fimbrial subunits, suggesting that it inhibits biofilm development by engaging the chaperone-usher machinery to disrupt polymer elongation. Functional DsbA is required for the selective packaging of DbaM into extracellular vesicles (EVs). Exogenously applied DbaM-EVs disrupted biofilms and altered APR resistance in the recipient cells. Collectively, these results define the Dsb-fimbriae-DbaM axis, linking periplasmic redox homeostasis, EV dynamics, and antibiotic susceptibility, which may offer potential targets for antimicrobial strategies.