Description of protein dynamics is known to be essential in understanding their function. Studies based on a well established \(^{15}\hbox {N}\) NMR relaxation methodology have been applied to a large number of systems. However, the low dispersion of \(^{1}\hbox {H}\) chemical shifts very often observed within intrinsically disordered proteins complicates utilization of standard 2D HN correlated spectra because a limited number of amino acids can be characterized. Here we present a suite of triple resonance HNCO-type NMR experiments for measurements of five \(^{15}\hbox {N}\) relaxation parameters ( \(R_1\) , \(R_2\) , NOE, cross-correlated relaxation rates \(\Gamma _x\) and \(\Gamma _z\) ) in doubly \(^{13}\hbox {C}\) , \(^{15}\hbox {N}\) -labeled proteins. We show that the third spectral dimension combined with non-uniform sampling provides relaxation rates for almost all residues of a protein with extremely poor chemical shift dispersion, the C terminal domain of \(\delta\) -subunit of RNA polymerase from Bacillus subtilis. Comparison with data obtained using a sample labeled by \(^{15}\hbox {N}\) only showed that the presence of \(^{13}\hbox {C}\) has a negligible effect on \(\Gamma _x\) , \(\Gamma _z\) , and on the cross-relaxation rate (calculated from NOE and \(R_1\) ), and that these relaxation rates can be used to calculate accurate spectral density values. Partially \(^{13}\hbox {C}\) -labeled sample was used to test if the observed increase of \(^{15}\hbox {N}\) \(R_1\) in the presence of \(^{13}\hbox {C}\) corresponds to the \(^{15}\hbox {N}-^{13}\hbox {C}\) dipoleâ??dipole interactions in the \(^{13}\hbox {C}\) , \(^{15}\hbox {N}\) -labeled sample.
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...
[NMR paper] Longitudinal relaxation properties of (1)H(N) and (1)H(?) determined by direct-detected (13)C NMR experiments to study intrinsically disordered proteins (IDPs).
Longitudinal relaxation properties of (1)H(N) and (1)H(?) determined by direct-detected (13)C NMR experiments to study intrinsically disordered proteins (IDPs).
Longitudinal relaxation properties of (1)H(N) and (1)H(?) determined by direct-detected (13)C NMR experiments to study intrinsically disordered proteins (IDPs).
J Magn Reson. 2015 Feb 12;254:19-26
Authors: Hošek T, Gil-Caballero S, Pierattelli R, Brutscher B, Felli IC
Abstract
Intrinsically disordered proteins (IDPs) are functional proteins containing large...
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03-17-2015 05:12 PM
Longitudinal relaxation properties of 1HN and 1H? determined by direct-detected 13C NMR experiments to study intrinsically disordered proteins (IDPs)
Longitudinal relaxation properties of 1HN and 1H? determined by direct-detected 13C NMR experiments to study intrinsically disordered proteins (IDPs)
Publication date: Available online 12 February 2015
Source:Journal of Magnetic Resonance</br>
Author(s): Tomáš Hošek , Sergi Gil-Caballero , Roberta Pierattelli , Bernhard Brutscher , Isabella C. Felli</br>
Intrinsically disordered proteins (IDPs) are functional proteins containing large fragments characterized by high local mobility. Bioinformatic studies have suggested that a significant fraction (more than 30%)...
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02-12-2015 07:48 PM
Triple resonance-based 13 C α and 13 C β CEST experiments for studies of ms timescale dynamics in proteins
Triple resonance-based 13 C α and 13 C β CEST experiments for studies of ms timescale dynamics in proteins
Abstract
A pair of triple resonance based CEST pulse schemes are presented for measuring 13Cα and 13Cβ chemical shifts of sparsely populated and transiently formed conformers that are invisible to traditional NMR experiments. CEST profiles containing dips at resonance positions of 13Cα or 13Cβ spins of major (ground) and minor (excited) conformers are obtained in a pseudo 3rd dimension that is generated by quantifying modulations of cross...
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10-28-2014 02:42 PM
New 13C-detected experiments for the assignment of intrinsically disordered proteins
New 13C-detected experiments for the assignment of intrinsically disordered proteins
Abstract
NMR assignment of intrinsically disordered proteins (IDPs) by conventional HN-detected methods is hampered by the small dispersion of the amide protons chemical shifts and exchange broadening of amide proton signals. Therefore several alternative assignment strategies have been proposed in the last years. Attempting to seize that dispersion of 13Câ?² and 15N chemical shifts holds even in IDPs, we recently proposed two 13C-detected experiments to directly...
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06-19-2014 10:21 PM
[NMR paper] High-dimensionality (13)C direct-detected NMR experiments for the automatic assignment of intrinsically disordered proteins.
High-dimensionality (13)C direct-detected NMR experiments for the automatic assignment of intrinsically disordered proteins.
Related Articles High-dimensionality (13)C direct-detected NMR experiments for the automatic assignment of intrinsically disordered proteins.
J Biomol NMR. 2013 Nov 8;
Authors: Bermel W, Felli IC, Gonnelli L, Ko?mi?ski W, Piai A, Pierattelli R, Zawadzka-Kazimierczuk A
Abstract
We present three novel exclusively heteronuclear 5D (13)C direct-detected NMR experiments, namely (H(N-flip)N)CONCACON, (HCA)CONCACON and...
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11-11-2013 01:30 AM
Extension of the HA-detection based approach: (HCA)CON(CA)H and (HCA)NCO(CA)H experiments for the main-chain assignment of intrinsically disordered proteins
Extension of the HA-detection based approach: (HCA)CON(CA)H and (HCA)NCO(CA)H experiments for the main-chain assignment of intrinsically disordered proteins
Abstract Extensive resonance overlap exacerbates assignment of intrinsically disordered proteins (IDPs). This issue can be circumvented by utilizing 15N, 13C� and 1HN spins, where the chemical shift dispersion is mainly dictated by the characteristics of consecutive amino acid residues. Especially 15N and 13C� spins offer superior chemical shift dispersion in comparison to 13Cα and 13Cβ spins. However, HN-detected experiments...
Triple resonance $$^{15}\hbox {N}$$ 15 N NMR relaxation experiments for studies of intrinsically disordered proteins
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