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Alamethicin Supramolecular Organization in Lipid Membranes from ¹⁹F Solid-State NMR.

ABSTRACT: Alamethicins (ALMs) are antimicrobial peptides of fungal origin. Their sequences are rich in hydrophobic amino acids and strongly interact with lipid membranes, where they cause a well-defined increase in conductivity. Therefore, the peptides are thought to form transmembrane helical bundles in which the more hydrophilic residues line a water-filled pore. Whereas the peptide has been well characterized in terms of secondary structure, membrane topology, and interactions, much fewer data are available regarding the quaternary arrangement of the helices within lipid bilayers. A new, to our knowledge, fluorine-labeled ALM derivative was prepared and characterized when reconstituted into phospholipid bilayers. As a part of these studies, C¹⁹F₃-labeled compounds were characterized and calibrated for the first time, to our knowledge, for ¹⁹F solid-state NMR distance and oligomerization measurements by centerband-only detection of exchange (CODEX) experiments, which opens up a large range of potential labeling schemes. The ¹⁹F-¹⁹F CODEX solid-state NMR experiments performed with ALM in POPC lipid bilayers and at peptide/lipid ratios of 1:13 are in excellent agreement with molecular-dynamics calculations of dynamic pentameric assemblies. When the peptide/lipid ratio was lowered to 1:30, ALM was found in the dimeric form, indicating that the supramolecular organization is tuned by equilibria that can be shifted by changes in environmental conditions.

SUBMITTER: Salnikov ES

PROVIDER: S-EPMC5153535 | biostudies-literature | 2016 Dec

REPOSITORIES: biostudies-literature

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Alamethicin Supramolecular Organization in Lipid Membranes from <sup>19</sup>F Solid-State NMR.

Salnikov Evgeniy S ES Raya Jesus J De Zotti Marta M Zaitseva Ekaterina E Peggion Cristina C Ballano Gema G Toniolo Claudio C Raap Jan J Bechinger Burkhard B

Biophysical journal 20161201 11

Alamethicins (ALMs) are antimicrobial peptides of fungal origin. Their sequences are rich in hydrophobic amino acids and strongly interact with lipid membranes, where they cause a well-defined increase in conductivity. Therefore, the peptides are thought to form transmembrane helical bundles in which the more hydrophilic residues line a water-filled pore. Whereas the peptide has been well characterized in terms of secondary structure, membrane topology, and interactions, much fewer data are avai ...[more]

PMID: 27926846

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Project description:Solid-state NMR spectra of membrane proteins often show significant line broadening at cryogenic temperatures. Here we investigate the effects of several cryoprotectants to preserve the spectral resolution of lipid membranes and membrane peptides at temperatures down to ~200 K. Trehalose, glycerol, dimethylsulfoxide (DMSO), dimethylformamide (DMF), and polyethylene glycol (PEG), were chosen. These compounds are commonly used in protein crystallography and cryobiology. 13C and 1H magic-angle-spinning spectra of several types of lipid membranes show that DMSO provides the best resolution enhancement over unprotected membranes and also best retards ice formation at low temperature. DMF and PEG-400 show slightly weaker cryoprotection, while glycerol and trehalose neither prevent membrane line broadening nor prevent ice formation under the conditions of our study. Neutral saturated-chain phospholipids are the most amenable to cryoprotection, whereas negatively charged and unsaturated lipids attenuate cryoprotection. 13C-1H dipolar couplings and 31P chemical shift anisotropies indicate that high spectral resolution at low temperature is correlated with stronger immobilization of the lipids at high temperature, indicating that line narrowing results from reduction of the conformational space sampled by the lipid molecules at high temperature. DMSO selectively narrowed the linewidths of the most disordered residues in the influenza M2 transmembrane peptide, while residues that exhibit narrow linewidths in the unprotected membrane are less impacted. A relatively rigid ?-hairpin antimicrobial peptide, PG-1, showed a linewidth increase of ~0.5 ppm over a ~70 K temperature drop both with and without cryoprotection. Finally, a short-chain saturated lipid, DLPE, exhibits excellent linewidths, suggesting that it may be a good medium for membrane protein structure determination. The three best cryoprotectants found in this work-DMSO, PEG, and DMF-should be useful for low-temperature membrane-protein structural studies by SSNMR without compromising spectral resolution.

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Alamethicin Supramolecular Organization in Lipid Membranes from ¹⁹F Solid-State NMR.

Publications

Alamethicin Supramolecular Organization in Lipid Membranes from <sup>19</sup>F Solid-State NMR.

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Alamethicin Supramolecular Organization in Lipid Membranes from 19F Solid-State NMR.

Publications

Alamethicin Supramolecular Organization in Lipid Membranes from <sup>19</sup>F Solid-State NMR.

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OmicsDI is part of the ELIXIR infrastructure

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Alamethicin Supramolecular Organization in Lipid Membranes from ¹⁹F Solid-State NMR.