Kanamycin uptake into Escherichia coli is facilitated by OmpF and OmpC porin channels located in the outer membrane

Despite decades of therapeutic application of aminoglycosides, it is still a matter of debate if porins contribute to the translocation of the antibiotics across the bacterial outer membrane. Here, we quantified the uptake of kanamycin across the major porin channels OmpF and OmpC present in the outer membrane of Escherichia coli. Our analysis revealed that, despite its relatively large size, about 10–20 kanamycin molecules per second permeate through OmpF and OmpC under a 10 μM concentration gradient, whereas OmpN does not allow the passage. Molecular simulations elucidate the uptake mechanism of kanamycin through these porins. Whole-cell studies with a defined set of E. coli porin mutants provide evidence that translocation of kanamycin via porins is relevant for antibiotic potency. The values are discussed with respect to other antibiotics

Ubiquitously expressed Human Beta Defensin 1 (hBD1) forms bacteria-entrapping nets in a redox dependent mode of action

Ever since the discovery of endogenous host defense antimicrobial peptides it has been discussed how these evolutionary conserved molecules avoid to induce resistance and to remain effective. Human beta-defensin 1 (hBD1) is an ubiquitously expressed endogenous antimicrobial peptide that exhibits qualitatively distinct activities between its oxidized and reduced forms. Here, we explore these antimicrobial mechanisms. Surprisingly, using electron microscopy we detected a so far unknown net-like structure surrounding bacteria, which were treated with the reduced but not the oxidized form of hBD1. A transmigration assay demonstrated that hBD1-derived nets capture bacteria and inhibit bacterial transmigration independent of bacterial killing. The presence of nets could completely prevent migration of hBD1 resistant pathogens and are stable in the presence of human duodenal secretion with a high amount of proteases. In contrast to HD6, cysteins are necessary for net formation. This redox-dependent function serves as an additional mechanism of action for hBD1 and differs from net formation by other defensins such as Paneth cell-derived human alpha-defensin 6 (HD6). While hBD1red and hBD1ox have distinct antimicrobial profiles and functions, only the reduced form provides additional host protection by entrapping bacteria in extracellular net structures preventing bacterial invasion. Better understanding of the modes of action of endogenous host peptides will help to find new antimicrobial strategies