Journal
INTERNATIONAL JOURNAL OF PEPTIDE RESEARCH AND THERAPEUTICS
Volume 15, Issue 2, Pages 147-155Publisher
SPRINGER
DOI: 10.1007/s10989-009-9182-3
Keywords
Antimicrobial peptides; Cathelicidin; Proline-rich peptides; Bac7; DnaK; Protein folding; Protein-protein interaction
Categories
Funding
- Italian Ministry for University and Research (PRIN 2007)
- Friuli Venezia Giulia Region [LR 26/2005]
- Stiftung fur Medizinische Forschung und Entwicklung, Zurich
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Bac7, a cathelicidin peptide of the proline-rich group, inactivates bacteria in a stereospecific manner by entering target cells without any apparent membrane damage and by binding to as yet unknown intracellular targets. The present study was aimed at detecting these putative intracellular interactors, which might mediate the antibacterial action of this peptide. By using affinity resins functionalized with the N-terminal 1-35 fragment of Bac7, a single protein was specifically retained with high affinity from Escherichia coli cytoplasmic protein lysates. This ligand was identified as the heat shock protein DnaK, the Hsp70 homolog in E. coli. The interaction between the peptide and the chaperone is stereospecific, given that a resin prepared with the all- d enantiomer failed to retain the protein. In vitro, Bac7(1-35) formed a complex with DnaK with an affinity comparable to that of other known high-affinity peptide ligands. In addition, at 10-100 mu M concentration, the peptide inhibited the protein refolding activity of the complete DnaK/DnaJ/GrpE/ATP molecular chaperone system in a dose-dependent manner. Despite these results, the in vitro sensitivity to the peptide, under growth permitting conditions, of DnaK-deficient E. coli strains was not significantly affected compared to the wild-type strain. This suggests that, apart from DnaK, other vital targets for the proline-rich AMPs are present in susceptible bacteria.
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