Conditions affecting the re-alignment of the antimicrobial peptide PGLa in membranes as monitored by solid state H-NMR |
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Authors: | Pierre Tremouilhac Parvesh Wadhwani |
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Institution: | a Institute for Biological Interfaces, Forschungszentrum Karlsruhe, Hermann-von-Helmholtz-Platz 1, 76344 Eggenstein-Leopoldshafen, Germany b Institute of Organic Chemistry, University of Karlsruhe, Fritz-Haber-Weg 6, 76131 Karlsruhe, Germany |
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Abstract: | The cationic antimicrobial peptide PGLa is electrostatically attracted to bacterial membranes, binds as an amphiphilic α-helix, and is thus able to permeabilize the lipid bilayer. Using solid state 2H-NMR of non-perturbing Ala-d3 labels on the peptide, we have characterized the helix alignment under a range of different conditions. Even at a very high peptide-to-lipid ratio (1:20) and in the presence of negatively charged lipids, there was no indication of a toroidal wormhole structure. Instead, PGLa re-aligns from a surface-bound S-state to an obliquely tilted T-state, which is presumably dimeric. An intermediate structure half-way between the S- and T-state was observed in fully hydrated multilamellar DMPC vesicles at 1:50, suggesting a fast exchange between the two states on the time scale of >50 kHz. We demonstrate that this equilibrium is shifted from the S- towards the T-state either upon (i) increasing the peptide concentration, (ii) adding negatively charged DMPG, or (iii) decreasing the level of hydration. The threshold concentration for re-alignment in DMPC is found to be between 1:200 and 1:100 in oriented samples at 96% humidity. In fully hydrated multilamellar DMPC vesicles, it shifts to an effective peptide-to-lipid ratio of 1:50 as some peptides are able to escape into the bulk water phase. |
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Keywords: | Ala-d3 3 3 3-2H3-l-alanine CD circular dichroism DMPC 1- 2-dimyristoyl-sn-glycero-3-phosphatidylcholine DMPG 1- 2-dimyristoyl-sn-glycero-3-phosphatidylglycerol MALDI-TOF matrix assisted laser desorption ionization-time of flight MLV multilamellar vesicles NMR nuclear magnetic resonance OCD oriented circular dichroism OS oriented sample PC phosphatidylcholine PG phosphatidylglycerol PGLa peptidyl-glycine-leucine-carboxyamide P/L peptide-to-lipid molar ratio RMSD root mean square deviation Smol molecular order parameter ΔνQ quadrupole splitting ρ peptide azimuthal rotation angle τ peptide tilt angle |
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