The Role of Bacterial Peptide Transporters in Intrinsic Resistance to Beta-lactam Antibiotics

Bacteria utilise peptides for a variety of biological functions. For bacteria to acquire peptides from the extracellular environment, they must pass the normally impermeable plasma membrane. For this purpose, a series of peptide transport proteins have evolved which facilitate the uptake of peptides into the cytoplasm. Despite peptide uptake being their primary function, peptide transporters have also been shown to facilitate the uptake of various drug molecules. One example is the beta-lactam antibiotics, the transport of which can be rationalised due to the presence of a peptide bond within their structure. The betalactam antibiotics exert their effect through binding to penicillin-binding proteins found within the periplasmic space, therefore the ability of peptide transporters to translocate beta-lactam antibiotics from the periplasmic space into the cytoplasm (and thus outside of their site of action) may contribute to the basal level of resistance to these drugs – a novel and previously untested hypothesis. These initial results described in this study document the creation by previous lab members of two mutant E. coli strains (known as ΔDB1 or ΔDB50) which lack either of the two peptide transporter classes present in E. coli (ABC and POT respectively). The comprehensive account of the creation of these strains, combined with confirmation of their genotype conducted in this work, allows these strains to now be used to investigate the role of these peptide transporters. To examine the role of peptide transporters in antibiotic resistance, growth of ΔDB50 was compared with that of wild type E. coli in the presence of various beta-lactam antibiotics with differing structural modifications. This revealed a consistently higher susceptibility in the mutant strain, an observation which was not present when exposed to antibiotics lacking a peptide bond, revealing a previously undescribed mechanism of antibiotic resistance. Phylogenetic analysis performed in this study demonstrates that POT transporters have significant diversity and are widely distributed across pathogenic bacterial species against which antibiotics are primarily used. Meanwhile, growth comparisons of ΔDB1 and ΔDB50 in the presence of aminoglycoside antibiotics revealed increased resistance in ΔDB1 but not ΔDB50, demonstrating a difference in antibiotic affinity of the two transporter types. Toxic peptide assays also revealed differences in transport specificity between the two classes depending on the stereoisomerism of the oligopeptide. The findings of this study therefore highlight the importance of understanding peptide transport in pathogenic bacteria and provide a potential insight into the nature of E. coli resistance to antibiotics which could be used to inform future strategies to combat this growing threat, alongside revealing additional differences in the transport specificities of peptide transporter classes.