Publication date: Available online 30 September 2014 Source:Journal of Magnetic Resonance
Author(s): Frank Löhr , Aisha Laguerre , Christoph Bock , Sina Reckel , Peter J. Connolly , Norzehan Abdul-Manan , Franz Tumulka , Rupert Abele , Jonathan M. Moore , Volker Dötsch
Combinatorial triple-selective labeling facilitates the NMR assignment process for proteins that are subject to signal overlap and insufficient signal-to-noise in standard triple-resonance experiments. Aiming at maximum amino-acid type and sequence-specific information, the method represents a trade-off between the number of selectively labeled samples that have to be prepared and the number of spectra to be recorded per sample. In order to address the demand of long measurement times, we here propose pulse sequences in which individual phase-shifted transients are stored separately and recombined later to produce several 2D HN(CX) type spectra that are usually acquired sequentially. Sign encoding by the phases of 13C 90° pulses allows to either select or discriminate against 13C’ or 13C? spins coupled to 15N. As a result, 1H-15N correlation maps of the various isotopomeric species present in triple-selectively labeled proteins are deconvoluted which in turn reduces problems due to spectral overlap. The new methods are demonstrated with four different membrane proteins with rotational correlation times ranging from 18 to 52 ns. Graphical abstract
[NMR paper] Improved NMR experiments with (13)C-isotropic mixing for assignment of aromatic and aliphatic side chains in labeled proteins.
Improved NMR experiments with (13)C-isotropic mixing for assignment of aromatic and aliphatic side chains in labeled proteins.
http://www.bionmr.com//www.ncbi.nlm.nih.gov/corehtml/query/egifs/http:--production.springer.de-OnlineResources-Logos-springerlink.gif Related Articles Improved NMR experiments with (13)C-isotropic mixing for assignment of aromatic and aliphatic side chains in labeled proteins.
J Biomol NMR. 2014 Jan 4;
Authors: Kovacs H, Gossert A
Abstract
Three improved (13)C-spinlock experiments for side chain assignments of...
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[NMR paper] Efficient resonance assignment of proteins in MAS NMR by simultaneous intra- and inter-residue 3D correlation spectroscopy.
Efficient resonance assignment of proteins in MAS NMR by simultaneous intra- and inter-residue 3D correlation spectroscopy.
http://www.ncbi.nlm.nih.gov/corehtml/query/egifs/http:--production.springer.de-OnlineResources-Logos-springerlink.gif Related Articles Efficient resonance assignment of proteins in MAS NMR by simultaneous intra- and inter-residue 3D correlation spectroscopy.
J Biomol NMR. 2013 Jan 19;
Authors: Daviso E, Eddy MT, Andreas LB, Griffin RG, Herzfeld J
Abstract
Resonance assignment is the first step in NMR structure...
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02-03-2013 10:19 AM
Multidimensional oriented solid-state NMR experiments enable the sequential assignment of uniformly 15N labeled integral membrane proteins in magnetically aligned lipid bilayers
Multidimensional oriented solid-state NMR experiments enable the sequential assignment of uniformly 15N labeled integral membrane proteins in magnetically aligned lipid bilayers
Abstract Oriented solid-state NMR is the most direct methodology to obtain the orientation of membrane proteins with respect to the lipid bilayer. The method consists of measuring 1H-15N dipolar couplings (DC) and 15N anisotropic chemical shifts (CSA) for membrane proteins that are uniformly aligned with respect to the membrane bilayer. A significant advantage of this approach is that tilt and azimuthal...
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10-10-2011 06:27 AM
Time-shared HSQC-NOESY for accurate distance constraints measured at high-field in 15N-13C-ILV methyl labeled proteins
Time-shared HSQC-NOESY for accurate distance constraints measured at high-field in 15N-13C-ILV methyl labeled proteins
Abstract We present a time-shared 3D HSQC-NOESY experiment that enables one to simultaneously record 13C- and 15N-dispersed spectra in Ile, Leu and Val (ILV) methyl-labeled samples. This experiment is designed to delineate the two spectra which would otherwise overlap with one another when acquired together. These spectra display nOe correlations in the detected proton dimension, i.e. with maximum resolution. This is in contrast to NOESY-HSQC types of experiments that...
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01-09-2011 12:46 PM
Sparsely-sampled High-resolution 4-D Experiments for Efficient Backbone Resonance Assignment of Disordered Proteins
Sparsely-sampled High-resolution 4-D Experiments for Efficient Backbone Resonance Assignment of Disordered Proteins
Publication year: 2011
Source: Journal of Magnetic Resonance, In Press, Accepted Manuscript, Available online 4 January 2011</br>
Jie, Wen , Jihui, Wu , Pei, Zhou</br>
Intrinsically disordered proteins (IDPs) play important roles in many critical cellular processes. Due to their limited chemical shift dispersion, IDPs often require four pairs of resonance connectivities (H?, C?, C? and CO) for establishing sequential backbone assignment. Because most conventional 4-D...
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01-05-2011 11:03 AM
Pseudo-4D triple resonance experiments to resolve HN overlap in the backbone assignment of unfolded proteins
Pseudo-4D triple resonance experiments to resolve HN overlap in the backbone assignment of unfolded proteins
Abstract The solution NMR resonance assignment of the protein backbone is most commonly carried out using triple resonance experiments that involve 15N and 1HN resonances. The assignment becomes problematic when there is resonance overlap of 15Nâ??1HN cross peaks. For such residues, one cannot unambiguously link the â??leftâ?? side of the NH root to the â??rightâ?? side, and the residues associated with such overlapping HN resonances remain often unassigned. Here we present a...
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[NMR paper] NMR resonance assignment of selectively labeled proteins by the use of paramagnetic l
NMR resonance assignment of selectively labeled proteins by the use of paramagnetic ligands.
Related Articles NMR resonance assignment of selectively labeled proteins by the use of paramagnetic ligands.
J Biomol NMR. 2004 Oct;30(2):205-10
Authors: Cutting B, Strauss A, Fendrich G, Manley PW, Jahnke W
Selective isotopic labeling of larger proteins greatly simplifies protein NMR spectra and reduces signal overlap, but selectively labeled proteins cannot be easily assigned since the sequential assignment method is not applicable. Here we describe...
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Simultaneous detection of amide and methyl correlations using a time shared NMR experiment: application to binding epitope mapping
Simultaneous detection of amide and methyl correlations using a time shared NMR experiment: application to binding epitope mapping
Peter Würtz, Olli Aitio, Maarit Hellman and Perttu Permi
Journal of Biomolecular NMR; 2007; 39(2) pp 97 - 105
Abstract:
Simultaneous recording of different NMR parameters is an efficient way to reduce the overall experimental time and speed up structural studies of biological macromolecules. This can especially be beneficial in the case of fast NMR-based drug screening applications or for collecting NOE restraints, where prohibitively long data collection...
Time-shared experiments for efficient assignment of triple-selectively labeled proteins
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