Antimicrobial and Membrane Disrupting Activities of a Peptide Derived from the Human Cathelicidin Antimicrobial Peptide LL37 |
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Authors: | Sathiah Thennarasu Anmin Tan Rajesh Penumatchu Deborah L Heyl |
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Institution: | † Department of Biophysics, University of Michigan, Ann Arbor, Michigan ‡ Department of Chemistry, University of Michigan, Ann Arbor, Michigan § Department of Biological and Materials Sciences, University of Michigan, Ann Arbor, Michigan ¶ Department of Chemistry, Eastern Michigan University, Ypsilanti, Michigan |
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Abstract: | A 21-residue peptide segment, LL7-27 (RKSKEKIGKEFKRIVQRIKDF), corresponding to residues 7-27 of the only human cathelicidin antimicrobial peptide, LL37, is shown to exhibit potent activity against microbes (particularly Gram-positive bacteria) but not against erythrocytes. The structure, membrane orientation, and target membrane selectivity of LL7-27 are characterized by differential scanning calorimetry, fluorescence, circular dichroism, and NMR experiments. An anilinonaphthalene-8-sulfonic acid uptake assay reveals two distinct modes of Escherichia coli outer membrane perturbation elicited by LL37 and LL7-27. The circular dichroism results show that conformational transitions are mediated by lipid-specific interactions in the case of LL7-27, unlike LL37. It folds into an α-helical conformation upon binding to anionic (but not zwitterionic) vesicles, and also does not induce dye leakage from zwitterionic lipid vesicles. Differential scanning calorimetry thermograms show that LL7-27 is completely integrated with DMPC/DMPG (3:1) liposomes, but induces peptide-rich and peptide-poor domains in DMPC liposomes. 15N NMR experiments on mechanically aligned lipid bilayers suggest that, like the full-length peptide LL37, the peptide LL7-27 is oriented close to the bilayer surface, indicating a carpet-type mechanism of action for the peptide. 31P NMR spectra obtained from POPC/POPG (3:1) bilayers containing LL7-27 show substantial disruption of the lipid bilayer structure and agree with the peptide's ability to induce dye leakage from POPC/POPG (3:1) vesicles. Cholesterol is shown to suppress peptide-induced disorder in the lipid bilayer structure. These results explain the susceptibility of bacteria and the resistance of erythrocytes to LL7-27, and may have implications for the design of membrane-selective therapeutic agents. |
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Keywords: | AMP antimicrobial peptide ANS anilinonaphthalene-8-sulfonic acid CD circular dichroism CP cross-polarization DSC differential scanning calorimetry MIC minimum inhibitory concentration MLV multilamellar vesicle NMR nuclear magnetic resonance PBS phosphate-buffered saline PISEMA polarization inversion spin exchange at the magic  angle POPC 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylcholine POPG 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylglycerol SUV small unilamellar vesicle DMPC 1 2-dimyristoyl-sn-glycero-3-phosphatidylcholine DMPG 1 2-dimyristoyl-glycero-3-phosphatidylglycerol |
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