Microcin PDI Inhibits Antibiotic-Resistant Strains of Escherichia coli and through a Mechanism of Membrane Disruption and Protection by Homotrimer Self-Immunity.

Microcin PDI (MccPDI), a class IIa microcin that is produced by strains 25 and 284, is known to inhibit foodborne pathogenic enterohemorrhagic serotypes O157:H7 and O26. Here we demonstrate that MccPDI can inhibit strains and isolates that are multidrug resistant, the latter including strains known to cause urinary tract infections in people and companion animals. Two exceptions out of 17 strains were identified. One of the two resistant isolates (AR0349) has a mutation in a critical amino acid residue that was identified in previous work as a requisite for the MccPDI precursor protein (McpM) to interact with outer membrane porin F (OmpF) on susceptible cells. The second resistant strain (MAD 96) had no mutations in , but it was PCR positive for two antimicrobial peptides, of which colicin Ia/Ib likely inhibits the MccPDI-producing strain during coculture. Recombinant McpM was still effective against strain MAD 96. In an assessment of how MccPDI affects susceptible strains, results from both an extracellular ATP assay and a nucleic acid staining assay were consistent with membrane damage, while the addition of 200- to 600-Da polyethylene glycol (PEG) to cocultures protected against MccPDI (>600-Da PEG did not provide protection). Further studies using a paraformaldehyde cross-linking experiment and a bacterial two-hybrid assay demonstrated that MccPDI immunity protein (McpI) forms a multimeric complex with itself and presumably protects the producer strain from within the periplasm through an unknown mechanism. Microcins represent potential alternatives to conventional antibiotics for human and veterinary medicine. For them to be applied in this manner, however, we need to better understand their spectrum of activity, how these proteins interact with susceptible cells, and how producer cells are protected against the antimicrobial properties of the microcins. For microcin PDI (MccPDI), we report that the spectrum of activity likely includes most strains due to a conserved binding motif found on an outer membrane protein. has this motif as well and is susceptible to MccPDI killing via damage to the bacterial membrane. Receptor specificity suggests that these proteins could be used without causing large-scale disruptions to a microbiota, but this also increases the likelihood that resistance can evolve via random mutations. As with conventional antibiotics, good stewardship will be needed to preserve the efficacy of microcins should they be deployed for clinical use.