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 ~200Â*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. 13Câ??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 β-hairpin antimicrobial peptide, PG-1, showed a linewidth increase of ~0.5Â*ppm over a ~70Â*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 workâ??DMSO, PEG, and DMFâ??should be useful for low-temperature membrane-protein structural studies by SSNMR without compromising spectral resolution.
[NMR paper] A 2H solid-state NMR study of lipid clustering by cationic antimicrobial and cell-penetrating peptides in model bacterial membranes.
A 2H solid-state NMR study of lipid clustering by cationic antimicrobial and cell-penetrating peptides in model bacterial membranes.
http://www.bionmr.com//www.ncbi.nlm.nih.gov/corehtml/query/egifs/http:--linkinghub.elsevier.com-ihub-images-cellhub.gif Related Articles A 2H solid-state NMR study of lipid clustering by cationic antimicrobial and cell-penetrating peptides in model bacterial membranes.
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Authors: Kwon B, Waring AJ, Hong M
Abstract
Domain formation in bacteria-mimetic membranes...
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[NMR paper] Investigation of the mechanism of action of novel amphipathic peptides: Insights from solid-state NMR studies of oriented lipid bilayers.
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Authors: Fillion M, Noël M, Lorin A, Voyer N, Auger M
Abstract
We have investigated in the present study the effect of both non-selective and selective cationic 14-mer peptides on the lipid orientation of DMPC bilayers...
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[NMR paper] High-resolution paramagnetically enhanced solid-state NMR spectroscopy of membrane proteins at fast magic angle spinning.
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J Biomol NMR. 2013 Dec 13;
Authors: Ward ME, Wang S, Krishnamurthy S, Hutchins H, Fey M, Brown LS, Ladizhansky V
Abstract
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Long-Term-Stable Ether-Lipid vs Conventional Ester-Lipid Bicelles in Oriented Solid-State NMR: Altered Structural Information in Studies of Antimicrobial Peptides.
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J Phys Chem B. 2011 Feb 10;
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Uniform isotope labeling of a eukaryotic seven-transmembrane helical protein in yeast enables high-resolution solid-state NMR studies in the lipid environment
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Abstract Overexpression of isotope-labeled multi-spanning eukaryotic membrane proteins for structural NMR studies is often challenging. On the one hand, difficulties with achieving proper folding, membrane insertion, and native-like post-translational modifications frequently disqualify bacterial expression systems. On the other hand, eukaryotic cell cultures can be prohibitively expensive. One of the viable alternatives,...
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Uniform isotope labeling of a eukaryotic seven-transmembrane helical protein in yeast enables high-resolution solid-state NMR studies in the lipid environment.
Uniform isotope labeling of a eukaryotic seven-transmembrane helical protein in yeast enables high-resolution solid-state NMR studies in the lipid environment.
Uniform isotope labeling of a eukaryotic seven-transmembrane helical protein in yeast enables high-resolution solid-state NMR studies in the lipid environment.
J Biomol NMR. 2011 Jan 19;
Authors: Fan Y, Shi L, Ladizhansky V, Brown LS
Overexpression of isotope-labeled multi-spanning eukaryotic membrane proteins for structural NMR studies is often challenging. On the one hand, difficulties...
Solid State NMR of membrane peptides and proteins
Solid State NMR of membrane peptides and proteins
Lecture notes on "Solid State NMR of membrane peptides and proteins" by Dr. SK Straus from Univ. of British Columbia
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Cryoprotection of lipid membranes for high-resolution solid-state NMR studies of membrane peptides and proteins at low temperature
Linear Mode
