Abstract
Solid-state NMR can provide insight into protein structure and dynamics at the atomic level without inherent protein size limitations. However, a major hurdle to studying large proteins by solid-state NMR spectroscopy is related to spectral complexity and resonance overlap, which increase with molecular weight and severely hamper the assignment process. Here we show the use of two sets of experiments that expand the tool kit of 1H-detected assignment approaches, and which correlate a given amide pair either to the two adjacent CO-CA pairs (4D hCOCANH/hCOCAcoNH), or to the amide 1H of the neighboring residue (3D HcocaNH/HcacoNH, which can be extended to up to 5D). The experiments are based on efficient coherence transfers between backbone atoms using INEPT transfers between carbons and cross-polarization for heteronuclear transfers. We exemplify the usefulness of these experiments with applications to assemblies of deuterated, fully amide-protonated proteins from ca. 20 to 60 kDa monomer, at MAS frequencies from ca. 40 to 55 kHz. These experiments will also be applicable to protonated proteins at higher MAS frequencies. We report the resonance assignment of a domain within the 50.4 kDa bacteriophage T5 tube protein pb6, and compare these to solution-state NMR assignments of the isolated domain in solution. This comparison reveals contacts of this domain to the core of the polymeric tail tube assembly.
PMID: 28792111 [PubMed - as supplied by publisher]
Five and four dimensional experiments for robust backbone resonance assignment of large intrinsically disordered proteins: application to Tau3x protein
Five and four dimensional experiments for robust backbone resonance assignment of large intrinsically disordered proteins: application to Tau3x protein
Abstract
New experiments dedicated for large IDPs backbone resonance assignment are presented. The most distinctive feature of all described techniques is the employment of MOCCA-XY16 mixing sequences to obtain effective magnetization transfers between carbonyl carbon backbone nuclei. The proposed 4 and 5 dimensional experiments provide a high dispersion of obtained signals making them suitable for use...
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07-20-2016 08:15 AM
[NMR paper] NMR Backbone Assignment of Large Proteins by Using (13) C? -Only Triple-Resonance Experiments.
NMR Backbone Assignment of Large Proteins by Using (13) C? -Only Triple-Resonance Experiments.
NMR Backbone Assignment of Large Proteins by Using (13) C? -Only Triple-Resonance Experiments.
Chemistry. 2016 Jun 8;
Authors: Wei Q, Chen J, Mi J, Zhang J, Ruan K, Wu J
Abstract
Nuclear magnetic resonance (NMR) is a powerful tool to interrogate protein structure and dynamics residue by residue. However, the prerequisite chemical-shift assignment remains a bottleneck for large proteins due to the fast relaxation and the frequency...
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06-09-2016 07:44 PM
BSH-CP based 3D solid-state NMR experiments for protein resonance assignment
BSH-CP based 3D solid-state NMR experiments for protein resonance assignment
Abstract
We have recently presented band-selective homonuclear cross-polarization (BSH-CP) as an efficient method for COâ??CA transfer in deuterated as well as protonated solid proteins. Here we show how the BSH-CP COâ??CA transfer block can be incorporated in a set of three-dimensional (3D) solid-state NMR (ssNMR) pulse schemes tailored for resonance assignment of proteins at high static magnetic fields and moderate magic-angle spinning rates. Due to the achieved...
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06-19-2014 10:21 PM
[NMR paper] BSH-CP based 3D solid-state NMR experiments for protein resonance assignment.
BSH-CP based 3D solid-state NMR experiments for protein resonance assignment.
Related Articles BSH-CP based 3D solid-state NMR experiments for protein resonance assignment.
J Biomol NMR. 2014 Mar 1;
Authors: Shi C, Fasshuber HK, Chevelkov V, Xiang S, Habenstein B, Vasa SK, Becker S, Lange A
Abstract
We have recently presented band-selective homonuclear cross-polarization (BSH-CP) as an efficient method for CO-CA transfer in deuterated as well as protonated solid proteins. Here we show how the BSH-CP CO-CA transfer block can be incorporated...
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03-04-2014 06:37 PM
[NMR paper] A Suite of Solid-State NMR Experiments for RNA Intranucleotide Resonance Assignment in a 21 kDa Protein-RNA Complex.
A Suite of Solid-State NMR Experiments for RNA Intranucleotide Resonance Assignment in a 21 kDa Protein-RNA Complex.
Related Articles A Suite of Solid-State NMR Experiments for RNA Intranucleotide Resonance Assignment in a 21 kDa Protein-RNA Complex.
Angew Chem Int Ed Engl. 2013 Jul 26;
Authors: Marchanka A, Simon B, Carlomagno T
Abstract
Intranucleotide resonance of the 26mer box C/D RNA in complex with the L7Ae protein were assigned by solid-state NMR (ssNMR; see picture) spectroscopy. This investigation opens the way for studying RNA in...
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07-31-2013 12:00 PM
[NMR paper] (13) C-Detected Through-Bond Correlation Experiments for Protein Resonance Assignment by Ultra-Fast MAS Solid-State NMR.
(13) C-Detected Through-Bond Correlation Experiments for Protein Resonance Assignment by Ultra-Fast MAS Solid-State NMR.
Related Articles (13) C-Detected Through-Bond Correlation Experiments for Protein Resonance Assignment by Ultra-Fast MAS Solid-State NMR.
Chemphyschem. 2013 Apr 15;
Authors: Barbet-Massin E, Pell AJ, Knight MJ, Webber AL, Felli IC, Pierattelli R, Emsley L, Lesage A, Pintacuda G
Abstract
We present two sequences which combine ((1) H,(15) N) and ((15) N,(13) C) selective cross-polarization steps with an efficient variant...
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04-17-2013 08:15 PM
Three-dimensional deuterium-carbon correlation experiments for high-resolution solid-state MAS NMR spectroscopy of large proteins
Three-dimensional deuterium-carbon correlation experiments for high-resolution solid-state MAS NMR spectroscopy of large proteins
Abstract Well-resolved 2Hâ??13C correlation spectra, reminiscent of 1Hâ??13C correlations, are obtained for perdeuterated ubiquitin and for perdeuterated outer-membrane protein G (OmpG) from E. coli by exploiting the favorable lifetime of 2H double-quantum (DQ) states. Sufficient signal-to-noise was achieved due to the short deuterium T 1, allowing for high repetition rates and enabling 3D experiments with a 2Hâ??13C transfer step in a reasonable time....
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11-01-2011 01:52 AM
Four-dimensional heteronuclear correlation experiments for chemical shift assignment of solid proteins
Four-dimensional heteronuclear correlation experiments for chemical shift assignment of solid proteins
W. Trent Franks, Kathryn D. Kloepper, Benjamin J. Wylie and Chad M. Rienstra
Journal of Biomolecular NMR; 2007; 39(2); pp 107 - 131
Abstract:
Chemical shift assignment is the first step in all established protocols for structure determination of uniformly labeled proteins by NMR. The explosive growth in recent years of magic-angle spinning (MAS) solid-state NMR (SSNMR) applications is largely attributable to improved methods for backbone and side-chain chemical shift correlation...
Solid-state NMR H-N-(C)-H and H-N-C-C 3D/4D correlation experiments for resonance assignment of large proteins.
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