Cyclic antimicrobial R-, W-rich peptides: the role of peptide structure and <Emphasis Type="Italic">E. coli</Emphasis> outer and inner membranes in activity and the mode of action |
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Authors: | Christof Junkes Richard D Harvey Kenneth D Bruce Rudolf Dölling Mojtaba Bagheri Margitta Dathe |
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Institution: | (1) Leibniz Institute of Molecular Pharmacology (FMP), Robert-Roessle-Str. 10, 13125 Berlin, Germany;(2) Institute of Pharmaceutical Science, King’s College London, 150 Stamford Street, London, SE1 9NH, UK;(3) Biosyntan GmbH, Robert-Roessle-Str. 10, 13125 Berlin, Germany; |
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Abstract: | This study compares the effect of cyclic R-, W-rich peptides with variations in amino acid sequences and sizes from 5 to 12
residues upon Gram negative and Gram positive bacteria as well as outer membrane-deficient and LPS mutant Escherichia coli (E.
coli) strains to analyze the structural determinants of peptide activity. Cyclo-RRRWFW (c-WFW) was the most active and E.
coli-selective sequence and bactericidal at the minimal inhibitory concentration (MIC). Removal of the outer membrane distinctly
reduced peptide activity and the complete smooth LPS was required for maximal activity. c-WFW efficiently permeabilised the
outer membrane of E.
coli and promoted outer membrane substrate transport. Isothermal titration calorimetric studies with lipid A-, rough-LPS (r-LPS)-
and smooth-LPS (s-LPS)-doped POPC liposomes demonstrated the decisive role of O-antigen and outer core polysaccharides for
peptide binding and partitioning. Peptide activity against the inner E. coli membrane (IM) was very low. Even at a peptide to lipid ratio of 8/1, c-WFW was not able to permeabilise a phosphatidylglycerol/phosphatidylethanolamine
(POPG/POPE) bilayer. Low influx of propidium iodide (PI) into bacteria confirmed a low permeabilising ability of c-WFW against
PE-rich membranes at the MIC. Whilst the peptide effect upon eukaryotic cells correlated with the amphipathicity and permeabilisation
of neutral phosphatidylcholine bilayers, suggesting a membrane disturbing mode of action, membrane permeabilisation does not
seem to be the dominating antimicrobial mechanism of c-WFW. Peptide interactions with the LPS sugar moieties certainly modulate
the transport across the outer membrane and are the basis of the E. coli selectivity of this type of peptides. |
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