Measuring the signs of the methyl 1 H chemical shift differences between major and â??invisibleâ?? minor protein conformational states using methyl 1 H multi-quantum spectroscopy
Measuring the signs of the methyl 1 H chemical shift differences between major and â??invisibleâ?? minor protein conformational states using methyl 1 H multi-quantum spectroscopy
Carrâ??Purcellâ??Meiboomâ??Gill (CPMG) type relaxation dispersion experiments are now routinely used to characterise protein conformational dynamics that occurs on the μs to millisecond (ms) timescale between a visible major state and â??invisibleâ?? minor states. The exchange rate(s) ( \( k_{{{\text{ex}}}} \) ), population(s) of the minor state(s) and the absolute value of the chemical shift difference \(|{\Delta \varpi }|\) (ppm) between different exchanging states can be extracted from the CPMG data. However the sign of \({\Delta \varpi }\) that is required to reconstruct the spectrum of the â??invisibleâ?? minor state(s) cannot be obtained from CPMG data alone. Building upon the recently developed triple quantum (TQ) methyl \( ^{1} {\text{H}} \) CPMG experiment (Yuwen in Angew Chem 55:11490â??11494, 2016) we have developed pulse sequences that use carbon detection to generate and evolve single quantum (SQ), double quantum (DQ) and TQ coherences from methyl protons in the indirect dimension to measure the chemical exchange-induced shifts of the SQ, DQ and TQ coherences from which the sign of \({\Delta \varpi }\) is readily obtained for two state exchange. Further a combined analysis of the CPMG data and the difference in exchange induced shifts between the SQ and DQ resonances and between the SQ and TQ resonances improves the estimates of exchange parameters like the population of the minor state. We demonstrate the use of these experiments on two proteins undergoing exchange: (1) the ~Â*18Â*kDa cavity mutant of T4 Lysozyme ( \( k_{{{\text{ex}}}} \sim\,3500{\text{ s}}^{{ - 1}} \) ) and (2) the \(\sim\,4.7\) Â*kDa Peripheral Sub-unit Binding Domain (PSBD) from the acetyl transferase of Bacillus stearothermophilus ( \(k_{ex} \sim\,13,000\hbox { s}^{-1}\) ).
An enhanced sensitivity methyl 1 H triple-quantum pulse scheme for measuring diffusion constants of macromolecules
An enhanced sensitivity methyl 1 H triple-quantum pulse scheme for measuring diffusion constants of macromolecules
Abstract
We present a pulse scheme that exploits methyl 1H triple-quantum (TQ) coherences for the measurement of diffusion rates of slowly diffusing molecules in solution. It is based on the well-known stimulated echo experiment, with encoding and decoding of TQ coherences. The size of quantifiable diffusion coefficients is thus lowered by an order of magnitude with respect to single-quantum (SQ) approaches. Notably, the sensitivity of...
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07-17-2017 04:07 PM
[NMR paper] A Conformational Ensemble Derived Using NMR Methyl Chemical Shifts Reveals a Mechanical Clamping Transition That Gates the Binding of the HU Protein to DNA.
A Conformational Ensemble Derived Using NMR Methyl Chemical Shifts Reveals a Mechanical Clamping Transition That Gates the Binding of the HU Protein to DNA.
Related Articles A Conformational Ensemble Derived Using NMR Methyl Chemical Shifts Reveals a Mechanical Clamping Transition That Gates the Binding of the HU Protein to DNA.
J Am Chem Soc. 2014 Feb 12;136(6):2204-7
Authors: Kannan A, Camilloni C, Sahakyan AB, Cavalli A, Vendruscolo M
Abstract
Recent improvements in the accuracy of structure-based methods for the prediction of...
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02-13-2014 03:35 PM
Long-lived nuclear spin states in methyl groups and quantum-rotor-induced polarization
From The DNP-NMR Blog:
Long-lived nuclear spin states in methyl groups and quantum-rotor-induced polarization
Meier, B., et al., Long-lived nuclear spin states in methyl groups and quantum-rotor-induced polarization. J Am Chem Soc, 2013. 135(50): p. 18746-9.
http://www.ncbi.nlm.nih.gov/pubmed/24252212
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02-13-2014 01:42 AM
Measurement of the signs of methyl 13C chemical shift differences between interconverting ground and excited protein states by R1Ï?: an application to αB-crystallin
Measurement of the signs of methyl 13C chemical shift differences between interconverting ground and excited protein states by R1Ï?: an application to αB-crystallin
Abstract Carr-Purcell-Meiboom-Gill relaxation dispersion (CPMG RD) NMR spectroscopy has emerged as a powerful tool for quantifying the kinetics and thermodynamics of millisecond time-scale exchange processes involving the interconversion between a visible ground state and one or more minor, sparsely populated invisible â??excitedâ?? conformational states. Recently it has also become possible to determine atomic resolution...
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04-09-2012 01:19 AM
Automated sequence- and stereo-specific assignment of methyl-labeled proteins by paramagnetic relaxation and methylâ??methyl nuclear overhauser enhancement spectroscopy
Automated sequence- and stereo-specific assignment of methyl-labeled proteins by paramagnetic relaxation and methylâ??methyl nuclear overhauser enhancement spectroscopy
Abstract Methyl-transverse relaxation optimized spectroscopy is rapidly becoming the preferred NMR technique for probing structure and dynamics of very large proteins up to ~1 MDa in molecular size. Data interpretation, however, necessitates assignment of methyl groups which still presents a very challenging and time-consuming process. Here we demonstrate that, in combination with a known 3D structure, paramagnetic...
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09-26-2011 06:42 AM
Backbone and Ile-?1, Leu, Val Methyl (1)H, (13)C and (15)N NMR chemical shift assignments for human interferon-stimulated gene 15 protein.
Backbone and Ile-?1, Leu, Val Methyl (1)H, (13)C and (15)N NMR chemical shift assignments for human interferon-stimulated gene 15 protein.
Backbone and Ile-?1, Leu, Val Methyl (1)H, (13)C and (15)N NMR chemical shift assignments for human interferon-stimulated gene 15 protein.
Biomol NMR Assign. 2011 May 5;
Authors: Yin C, Aramini JM, Ma LC, Cort JR, Swapna GV, Krug RM, Montelione GT
Human interferon-stimulated gene 15 protein (ISG15), also called ubiquitin cross-reactive protein (UCRP), is the first identified ubiquitin-like protein containing...
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05-06-2011 12:02 PM
A simple method for measuring signs of 1HN chemical shift differences between ground
Abstract NMR relaxation dispersion spectroscopy is a powerful method for studying protein conformational dynamics whereby visible, ground and invisible, excited conformers interconvert on the millisecond time-scale. In addition to providing kinetics and thermodynamics parameters of the exchange process, the CPMG dispersion experiment also allows extraction of the absolute values of the chemical shift differences between interconverting states,
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ê , opening the way for structure determination of excited state conformers. Central to the goal of structural...
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08-14-2010 04:19 AM
Measurement of signs of chemical shift differences between ground and excited protein
Abstract Carr-Purcell-Meiboom-Gill (CPMG) relaxation dispersion NMR spectroscopy has emerged as a powerful tool for quantifying the kinetics and thermodynamics of millisecond exchange processes between a major, populated ground state and one or more minor, low populated and often invisible â??excitedâ?? conformers. Analysis of CPMG data-sets also provides the magnitudes of the chemical shift difference(s) between exchanging states (|Î?Ï?|), that inform on the structural properties of the excited state(s). The sign of Î?Ï? is, however, not available from CPMG data. Here we present...
Measuring the signs of the methyl 1 H chemical shift differences between major and â??invisibleâ?? minor protein conformational states using methyl 1 H multi-quantum spectroscopy
Linear Mode
