18 research outputs found
Integrated evolutionary analysis reveals antimicrobial peptides with limited resistance
Antimicrobial peptides (AMPs) are promising antimicrobials, however, the potential of bacterial resistance is a major concern. Here we systematically study the evolution of resistance to 14 chemically diverse AMPs and 12 antibiotics in Escherichia coli. Our work indicates that evolution of resistance against certain AMPs, such as tachyplesin II and cecropin P1, is limited. Resistance level provided by point mutations and gene amplification is very low and antibiotic-resistant bacteria display no cross-resistance to these AMPs. Moreover, genomic fragments derived from a wide range of soil bacteria confer no detectable resistance against these AMPs when introduced into native host bacteria on plasmids. We have found that simple physicochemical features dictate bacterial propensity to evolve resistance against AMPs. Our work could serve as a promising source for the development of new AMP-based therapeutics less prone to resistance, a feature necessary to avoid any possible interference with our innate immune system
Rapid evolution of reduced susceptibility against a balanced dual targeting antibiotic through stepping-stone mutations
Multi-targeting antibiotics, i.e., single compounds capable of inhibiting two or more bacterial targets
are generally considered as a promising therapeutic strategy against resistance evolution. The
rationale for this theory is that multi-targeting antibiotics demand the simultaneous acquisition of
multiple mutations at their respective target genes to achieve significant resistance. The theory
presumes that individual mutations provide little or no benefit to the bacterial host. Here we propose
that such individual, stepping-stone mutations can be prevalent in clinical bacterial isolates, as they
provide significant resistance to other antimicrobial agents. To test this possibility, we focused on
gepotidacin, an antibiotic candidate that selectively inhibits both bacterial DNA gyrase and
topoisomerase IV. In a susceptible organism, Klebsiella pneumoniae, a combination of two specific
mutations in these target proteins provide an over 2000-fold reduction in susceptibility, while
individually none of these mutations affect resistance significantly. Alarmingly, strains with decreased
susceptibility against gepotidacin are found to be as virulent as the wild-type Klebsiella pneumoniae
strain in a murine model. Moreover, numerous pathogenic isolates carry mutations which could
promote the evolution of clinically significant reduction of susceptibility against gepotidacin in the
future. As might be expected, prolonged exposure to ciprofloxacin, a clinically widely employed
gyrase inhibitor, co-selected for reduced susceptibility against gepotidacin. We conclude that
extensive antibiotic usage could select for mutations that serve as stepping-stones towards resistance
against antimicrobial compounds still under development. Our research indicates that even balanced
multi-targeting antibiotics are prone to resistance evolution
Cotranslational protein assembly imposes evolutionary constraints on homomeric proteins
Cotranslational protein folding can facilitate rapid formation of functional structures. However, it might also cause premature assembly of protein complexes, if two interacting nascent chains are in close proximity. By analyzing known protein structures, we show that homomeric protein contacts are enriched towards the C-termini of polypeptide chains across diverse proteomes. We hypothesize that this is the result of evolutionary constraints for folding to occur prior to assembly. Using high-throughput imaging of protein homomers in vivo in E. coli and engineered protein constructs with N- and C-terminal oligomerization domains, we show that, indeed, proteins with C-terminal homomeric interface residues consistently assemble more efficiently than those with N-terminal interface residues. Using in vivo, in vitro and in silico experiments, we identify features that govern successful assembly of homomers, which have implications for protein design and expression optimization
Dual Action of the PN159/KLAL/MAP Peptide
The absorption of drugs is limited by the epithelial barriers of the gastrointestinal tract. One of the strategies to improve drug delivery is the modulation of barrier function by the targeted opening of epithelial tight junctions. In our previous study the 18-mer amphiphilic PN159 peptide was found to be an effective tight junction modulator on intestinal epithelial and blood⁻brain barrier models. PN159, also known as KLAL or MAP, was described to interact with biological membranes as a cell-penetrating peptide. In the present work we demonstrated that the PN159 peptide as a penetration enhancer has a dual action on intestinal epithelial cells. The peptide safely and reversibly enhanced the permeability of Caco-2 monolayers by opening the intercellular junctions. The penetration of dextran molecules with different size and four efflux pump substrate drugs was increased several folds. We identified claudin-4 and -7 junctional proteins by docking studies as potential binding partners and targets of PN159 in the opening of the paracellular pathway. In addition to the tight junction modulator action, the peptide showed cell membrane permeabilizing and antimicrobial effects. This dual action is not general for cell-penetrating peptides (CPPs), since the other three CPPs tested did not show barrier opening effects