Cryoprotection of lipid membranes for high-resolution solid-state NMR studies of membrane peptides and proteins at low temperature.


: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 β-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.


J Biomol NMR


Lee M,Hong M




Has Abstract


2014-08-01 00:00:00












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    journal_title:Journal of biomolecular NMR

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    authors: Hu KN,Qiang W,Tycko R

    更新日期:2011-07-01 00:00:00

  • Structural features of the metal binding site and dynamics of gallium putidaredoxin, a diamagnetic derivative of a Cys4Fe2S2 ferredoxin.

    abstract::The first reconstitution of an Fe2S2 ferredoxin with a diamagnetic prosthetic group was recently described [Kazanis et al. (1995) J. Am. Chem. Soc., 117, 6625-6626]. The replacement of the iron-sulfur cluster of the bacterial ferredoxin putidaredoxin (Pdx) by gallium (Ga3+) renders the protein diamagnetic and permits ...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章


    authors: Kazanis S,Pochapsky TC

    更新日期:1997-06-01 00:00:00

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    abstract::A simple triple resonance NMR experiment that leads to the correlation of the backbone amide resonances of each amino acid residue 'i' with that of residues 'i-1' and 'i+1' in ((13)C, (15)N) labelled intrinsically disordered proteins (IDPs) is presented. The experimental scheme, {HN-NCA heteronuclear TOCSY-NH}, exploi...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章


    authors: Wiedemann C,Goradia N,Häfner S,Herbst C,Görlach M,Ohlenschläger O,Ramachandran R

    更新日期:2015-10-01 00:00:00

  • 1H nuclear magnetic resonance determination of the membrane-bound conformation of senktide, a highly selective neurokinin B agonist.

    abstract::Senktide is a highly specific and potent analog of neurokinin B, the natural ligand of the tachykinin receptor NK-3. The membrane-bound conformation of senktide, interacting with negatively charged membrane vesicles composed of perdeuterated phosphatidylcholine and phosphatidylglycerol (70:30), has been investigated u...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章


    authors: Bersch B,Koehl P,Nakatani Y,Ourisson G,Milon A

    更新日期:1993-07-01 00:00:00

  • Improved 3D gd-HCACO and gd-(H)CACO-TOCSY experiments for isotopically enriched proteins dissolved in H2O.

    abstract::Pulsed field gradients were incorporated into the HCACO experiment for acquiring spectra on isotopically enriched protein samples dissolved in H2O. Excellent water suppression and spectral quality were achieved using the modified pulse sequence (gd-HCACO), as demonstrated for a 13C-/15N-labeled sample of the SH2 domai...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章


    authors: Zhang W,Gmeiner WH

    更新日期:1996-05-01 00:00:00

  • [13C]-constant-time [15N,1H]-TROSY-HNCA for sequential assignments of large proteins.

    abstract::The greatly improved sensitivity resulting from the use of TROSY during 15N evolution and amide proton acquisition enables the recording of HNCA spectra of large proteins with constant-time 13C alpha evolution. In [13C]-ct-[15N,1H]-TROSY-HNCA experiments with a 2H/13C/15N-labeled 110 kDa protein, 7,8-dihydroneopterin ...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章


    authors: Salzmann M,Pervushin K,Wider G,Senn H,Wüthrich K

    更新日期:1999-05-01 00:00:00

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    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章


    authors: Eccles C,Güntert P,Billeter M,Wüthrich K

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  • Resolving complex mixtures: trilinear diffusion data.

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    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章


    authors: Björnerås J,Botana A,Morris GA,Nilsson M

    更新日期:2014-04-01 00:00:00

  • Intraresidue 1H-15N-13C' and 1H alpha-13C alpha-13C' dipole-CSA relaxation interference as a source of constraints for structural refinement of metal-binding sites in zinc-finger proteins.

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    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章


    authors: Kloiber K,Schüler W,Konrat R

    更新日期:2001-04-01 00:00:00

  • Elucidation of the origin of multiple conformations of the human alpha 3-chain type VI collagen C-terminal Kunitz domain: the reorientation of the Trp21 ring.

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    journal_title:Journal of biomolecular NMR

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    authors: Sørensen MD,Kristensen SM,Bjørn S,Norris K,Olsen O,Led JJ

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    abstract::Active bandwidth and global quality factor are the two main metrics used to quantitatively compare the performance of TOCSY mixing sequences. Active bandwidth refers to the spectral region over which at least 50 % of the magnetization is transferred via a coupling. Global quality factor scores mixing sequences accordi...

    journal_title:Journal of biomolecular NMR

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    authors: Coote P,Bermel W,Wagner G,Arthanari H

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    abstract::In magic angle spinning solid state NMR experiments the potential of heteronuclear (1)H decoupling employing a continuous train of adiabatic inversion pulses has been assessed via numerical simulations and experimental measurements. It is shown that, with a (1)H RF field strength of approximately 100 kHz that is typic...

    journal_title:Journal of biomolecular NMR

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    authors: Leppert J,Ohlenschläger O,Görlach M,Ramachandran R

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    journal_title:Journal of biomolecular NMR

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    authors: Lange OF

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    journal_title:Journal of biomolecular NMR

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    authors: Nabuurs SB,Krieger E,Spronk CA,Nederveen AJ,Vriend G,Vuister GW

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    journal_title:Journal of biomolecular NMR

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    authors: Mueller GA

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    journal_title:Journal of biomolecular NMR

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    authors: Movellan KT,Najbauer EE,Pratihar S,Salvi M,Giller K,Becker S,Andreas LB

    更新日期:2019-02-01 00:00:00

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    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章


    authors: Hare BJ,Prestegard JH

    更新日期:1994-01-01 00:00:00

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    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章


    authors: Wishart DS,Wang Y

    更新日期:1998-04-01 00:00:00

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    journal_title:Journal of biomolecular NMR

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    authors: Sakhrani VV,Hilario E,Caulkins BG,Hatcher-Skeers ME,Fan L,Dunn MF,Mueller LJ

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    journal_title:Journal of biomolecular NMR

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    authors: Zweckstetter M

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    abstract::NMR relaxometry plays crucial role in studies of protein dynamics. The measurement of longitudinal and transverse relaxation rates of [Formula: see text]N is the main source of information on backbone motions. However, even the most basic approach exploiting a series of [Formula: see text]N HSQC spectra can require se...

    journal_title:Journal of biomolecular NMR

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    authors: Urbańczyk M,Nowakowski M,Koźmiński W,Kazimierczuk K

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    journal_title:Journal of biomolecular NMR

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    authors: Sternberg U,Witter R

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  • 1H(C) and 1H(N) total NOE correlations in a single 3D NMR experiment. 15N and 13C time-sharing in t1 and t2 dimensions for simultaneous data acquisition.

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    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章


    authors: Xia Y,Yee A,Arrowsmith CH,Gao X

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  • Spectral densities of nitrogen nuclei in Escherichia coli ribonuclease HI obtained by 15N NMR relaxation and molecular dynamics.

    abstract::Spectral densities of the 15N amide in Escherichia coli ribonuclease HI, obtained from NMR relaxation experiments, were compared with those calculated using a molecular dynamics (MD) simulation. All calculations and comparisons assumed that the auto-correlation function describing the internal motions of the molecule ...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章


    authors: Ishima R,Yamasaki K,Saito M,Nagayama K

    更新日期:1995-09-01 00:00:00

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    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章


    authors: Keiffer S,Carneiro MG,Hollander J,Kobayashi M,Pogoryelev D,Ab E,Theisgen S,Müller G,Siegal G

    更新日期:2020-11-01 00:00:00

  • Comprehensive analysis of NMR data using advanced line shape fitting.

    abstract::NMR spectroscopy is uniquely suited for atomic resolution studies of biomolecules such as proteins, nucleic acids and metabolites, since detailed information on structure and dynamics are encoded in positions and line shapes of peaks in NMR spectra. Unfortunately, accurate determination of these parameters is often co...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章


    authors: Niklasson M,Otten R,Ahlner A,Andresen C,Schlagnitweit J,Petzold K,Lundström P

    更新日期:2017-10-01 00:00:00

  • Trans-hydrogen bond deuterium isotope effects of A:T base pairs in DNA.

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    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章


    authors: Vakonakis I,LiWang AC

    更新日期:2004-05-01 00:00:00

  • Improved NMR spectra of a protein-DNA complex through rational mutagenesis and the application of a sensitivity optimized isotope-filtered NOESY experiment.

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    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章


    authors: Iwahara J,Wojciak JM,Clubb RT

    更新日期:2001-03-01 00:00:00

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    abstract::Two-dimensional (2D) 1H NMR experiments using deuterium labeling have been carried out to investigate the solution of ribonuclease HI (RNase HI) from Escherichia coli (E. coli), which consists of 155 amino acids. To simplify the 1H NMR spectra, two fully deuterated enzymes bearing several protonated amino acids were p...

    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章


    authors: Oda Y,Nakamura H,Yamazaki T,Nagayama K,Yoshida M,Kanaya S,Ikehara M

    更新日期:1992-03-01 00:00:00

  • Structure refinement of flexible proteins using dipolar couplings: application to the protein p8MTCP1.

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    journal_title:Journal of biomolecular NMR

    pub_type: 杂志文章


    authors: Déméné H,Ducat T,Barthe P,Delsuc MA,Roumestand C

    更新日期:2002-01-01 00:00:00