Recently, 15N-detected multidimensional NMR experiments have been introduced for the investigation of proteins. Utilization of the slow transverse relaxation of nitrogen nuclei in a 15N-TROSY experiment allowed recording of high quality spectra for high molecular weight proteins, even in the absence of deuteration. Here, we demonstrate the applicability of three 15N-detected Hâ??N correlation experiments (TROSY, BEST-TROSY and HSQC) to RNA. With the newly established 15N-detected BEST-TROSY experiment, which proves to be the most sensitive 15N-detected Hâ??N correlation experiment, spectra for five RNA molecules ranging in size from 5 to 100Â*kDa were recorded. These spectra yielded high resolution in the 15N-dimension even for larger RNAs since the increase in line width with molecular weight is more pronounced in the 1H- than in the 15N-dimension. Further, we could experimentally validate the difference in relaxation behavior of imino groups in AU and GC base pairs. Additionally, we showed that 15N-detected experiments theoretically should benefit from sensitivity and resolution advantages at higher static fields but that the latter is obscured by exchange dynamics within the RNAs.
[NMR paper] Solid-state NMR H-N-(C)-H and H-N-C-C 3D/4D correlation experiments for resonance assignment of large proteins.
Solid-state NMR H-N-(C)-H and H-N-C-C 3D/4D correlation experiments for resonance assignment of large proteins.
Related Articles Solid-state NMR H-N-(C)-H and H-N-C-C 3D/4D correlation experiments for resonance assignment of large proteins.
Chemphyschem. 2017 Aug 09;:
Authors: Fraga H, Arnaud CA, Gauto DF, Audin MJC, Kurauskas V, Macek P, Krichel C, Guan JY, Boisbouvier J, Sprangers R, Breyton C, Schanda P
Abstract
Solid-state NMR can provide insight into protein structure and dynamics at the atomic level without inherent protein...
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08-10-2017 01:27 PM
Nitrogen-detected TROSY yields comparable sensitivity to proton-detected TROSY for non-deuterated, large proteins under physiological salt conditions
Nitrogen-detected TROSY yields comparable sensitivity to proton-detected TROSY for non-deuterated, large proteins under physiological salt conditions
Abstract
Direct detection of the TROSY component of proton-attached 15N nuclei (15N-detected TROSY) yields high quality spectra with high field magnets, by taking advantage of the slow 15N transverse relaxation. The slow transverse relaxation and narrow line width of the 15N-detected TROSY resonances are expected to compensate for the inherently low 15N sensitivity. However, the sensitivity of...
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01-23-2016 03:35 PM
19 F-labeling of the adenine H2-site to study large RNAs by NMR spectroscopy
19 F-labeling of the adenine H2-site to study large RNAs by NMR spectroscopy
Abstract
In comparison to proteins and protein complexes, the size of RNA amenable to NMR studies is limited despite the development of new isotopic labeling strategies including deuteration and ligation of differentially labeled RNAs. Due to the restricted chemical shift dispersion in only four different nucleotides spectral resolution remains limited in larger RNAs. Labeling RNAs with the NMR-active nucleus 19F has previously been introduced for small RNAs up to 40...
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12-28-2015 12:26 AM
[NMR paper] (19)F-labeling of the adenine H2-site to study large RNAs by NMR spectroscopy.
(19)F-labeling of the adenine H2-site to study large RNAs by NMR spectroscopy.
Related Articles (19)F-labeling of the adenine H2-site to study large RNAs by NMR spectroscopy.
J Biomol NMR. 2015 Dec 24;
Authors: Sochor F, Silvers R, Müller D, Richter C, Fürtig B, Schwalbe H
Abstract
In comparison to proteins and protein complexes, the size of RNA amenable to NMR studies is limited despite the development of new isotopic labeling strategies including deuteration and ligation of differentially labeled RNAs. Due to the restricted...
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12-28-2015 12:26 AM
[NMR paper] Combining NMR and EPR to Determine Structures of Large RNAs and Protein-RNA Complexes in Solution.
Combining NMR and EPR to Determine Structures of Large RNAs and Protein-RNA Complexes in Solution.
Related Articles Combining NMR and EPR to Determine Structures of Large RNAs and Protein-RNA Complexes in Solution.
Methods Enzymol. 2015;558:279-331
Authors: Duss O, Yulikov M, Allain FH, Jeschke G
Abstract
Although functional significance of large noncoding RNAs and their complexes with proteins is well recognized, structural information for this class of systems is very scarce. Their inherent flexibility causes problems in...
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06-13-2015 11:09 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
Nitrogen-detected CAN and CON experiments as alternative experiments for main chain N
Abstract Heteronuclear direct-detection experiments, which utilize the slower relaxation properties of low γ nuclei, such as 13C have recently been proposed for sequence-specific assignment and structural analyses of large, unstructured, and/or paramagnetic proteins. Here we present two novel 15N direct-detection experiments. The CAN experiment sequentially connects amide 15N resonances using 13Cα chemical shift matching, and the CON experiment connects the preceding 13C� nuclei. When starting from the same carbon polarization, the intensities of nitrogen signals detected in the CAN or...
Evaluation of 15 N-detected Hâ??N correlation experiments on increasingly large RNAs
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