Chimeric Antimicrobial Peptides Exhibit Multiple Modes of Action

  • Laszlo OtvosJr.
  • Christine Snyder
  • Barry Condie
  • Philippe Bulet
  • John D. Wade
Open Access
Article

Abstract

Pyrrhocoricin and drosocin, representatives of the short, proline-rich antimicrobial peptide family kill bacteria by inactivating the bacterial heat shock protein DnaK and inhibiting chaperone-assisted protein folding. The molecular architecture of these peptides features an N-terminal DnaK-binding half and a C-terminal delivery unit, capable of crossing bacterial membranes. Cell penetration is enhanced if multiple copies of pyrrhocoricin are conjugated. To obtain drug leads with improved antimicrobial properties, and possible utility as therapeutic agents, we synthesized chimeric dimers, in which pyrrhocoricin’s potent DnaK-binding domain was connected to drosocin’s superior cell penetrating module. Indeed, the new constructs not only exhibited enhanced in vitro antibacterial properties against the originally sensitive strains Escherichia coli, Klebsiella pneumoniae and Salmonella typhimurium, but also showed activity against Staphylococcus aureus, a bacterial strain resistant to native pyrrhocoricin and drosocin. The improved antimicrobial profile could be demonstrated with assays designed to distinguish intracellular or membrane activities. While a novel mixed pyrrhocoricin–drosocin dimer and the purely pyrrhocoricin-based old dimer bound E. coli DnaK with an identical 4 μM K d, the mixed dimers penetrated a significantly larger number of E. coli and S. aureus cells than the previous analogs and destroyed a larger percentage of bacterial membrane structures. Toxicity to human red blood cells could not be observed up to the highest peptide concentration tested, 640 μM. In addition, repetitive reculturing of E. coli or S. aureus cells with sublethal concentrations of the mixed dimer did not result in resistance induction to the novel peptide antibiotic. The new concept of pyrrhocoricin–drosocin mixed dimers yields antibacterial peptide derivatives acting with a multiple mode of action, and can serve as a useful addition to the current antimicrobial therapy repertoire.

Key words

Cell penetration DnaK binding drosocin growth inhibition membrane disintegration pyrrhocoricin resistant bacteria stability toxicity 

Abbreviations

Chex

1-amino-1-cyclohexane carboxylic acid

Dab

2,4-diamino-butyric acid

DIC

differential interference contrast

Hsp

heat shock protein

LPS

lipopolysaccharide

MALDI

matrix-assisted laser desorption/ionization

MHB

Muller–Hinton broth

MIC

minimal inhibitory concentration

PBS

phosphate buffered saline

Pip

4-amino-piperidine-4-carboxylic acid

RP-HPLC

reversed-phase high performance liquid chromatography

TNF

tumor necrosis factor

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Copyright information

© Springer Science+Business Media, Inc. 2005

Authors and Affiliations

  • Laszlo OtvosJr.
    • 1
  • Christine Snyder
    • 1
  • Barry Condie
    • 1
  • Philippe Bulet
    • 2
  • John D. Wade
    • 3
  1. 1.The Wistar InstitutePhiladelphiaUSA
  2. 2.Atheris LaboratoriesGenevaSwitzerland
  3. 3.Howard Florey InstituteMelbourneAustralia

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