Publication date: Available online 10 May 2016 Source:Journal of Magnetic Resonance
Author(s): Zhongyu Yang, Michael D. Bridges, Carlos J. López, Olga Yu. Rogozhnikova, Dmitry V. Trukhin, Evan K. Brooks, Victor Tormyshev, Howard J. Halpern, Wayne L. Hubbell
Site-directed spin labeling (SDSL) in combination with Electron Paramagnetic Resonance (EPR) spectroscopy has become an important tool for measuring distances in proteins on the order of a few nm. For this purpose pairs of spin labels, most commonly nitroxides, are site-selectively introduced into the protein. Recent efforts to develop new spin labels are focused on tailoring the intrinsic properties of the label to either extend the upper limit of measurable distances at physiological temperature, or to provide a unique spectral lineshape so that selective pairwise distances can be measured in a protein or complex containing multiple spin label species. Triarylmethyl (TAM) radicals are the foundation for a new class of spin labels that promise to provide both capabilities. Here we report a new methanethiosulfonate derivative of a TAM radical that reacts rapidly and selectively with an engineered cysteine residue to generate a TAM containing side chain (TAM1) in high yield. With a TAM1 residue and Cu2+ bound to an engineered Cu2+ binding site, enhanced T1 relaxation of TAM should enable measurement of interspin distances up to 50 Å at physiological temperature. To achieve favorable TAM1-labeled protein concentrations without aggregation, proteins are tethered to a solid support either site-selectively using an unnatural amino acid or via native lysine residues. The methodology is general and readily extendable to complex systems, including membrane proteins. Graphical abstract
[NMR paper] Solution NMR Structure Determination of Polytopic ?-Helical Membrane Proteins: A Guide to Spin Label Paramagnetic Relaxation Enhancement Restraints.
Solution NMR Structure Determination of Polytopic ?-Helical Membrane Proteins: A Guide to Spin Label Paramagnetic Relaxation Enhancement Restraints.
http://www.bionmr.com//www.ncbi.nlm.nih.gov/corehtml/query/egifs/http:--linkinghub.elsevier.com-ihub-images-PubMedLink.gif Related Articles Solution NMR Structure Determination of Polytopic ?-Helical Membrane Proteins: A Guide to Spin Label Paramagnetic Relaxation Enhancement Restraints.
Methods Enzymol. 2015;557:329-348
Authors: Columbus L, Kroncke B
Abstract
Solution nuclear...
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[NMR paper] Relaxation-compensated difference spin diffusion NMR for detecting (13)C- (13)C long-range correlations in proteins and polysaccharides.
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J Biomol NMR. 2014 Dec 16;
Authors: Wang T, Williams JK, Schmidt-Rohr K, Hong M
Abstract
The measurement of long-range distances remains a challenge in solid-state NMR structure determination of biological macromolecules. In 2D and 3D correlation spectra of...
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Relaxation-compensated difference spin diffusion NMR for detecting 13 Câ?? 13 C long-range correlations in proteins and polysaccharides
Relaxation-compensated difference spin diffusion NMR for detecting 13 Câ?? 13 C long-range correlations in proteins and polysaccharides
Abstract
The measurement of long-range distances remains a challenge in solid-state NMR structure determination of biological macromolecules. In 2D and 3D correlation spectra of uniformly 13C-labeled biomolecules, inter-residue, inter-segmental, and intermolecular 13Câ??13C cross peaks that provide important long-range distance constraints for three-dimensional structures often overlap with short-range cross peaks...
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The Spatial Effect of Protein Deuteration on Nitroxide Spin-label Relaxation: Implications for EPR Distance Measurement
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Publication date: Available online 28 September 2014
Source:Journal of Magnetic Resonance</br>
Author(s): Hassane. El Mkami , Richard Ward , Andrew Bowman , Tom Owen-Hughes , David G. Norman</br>
Pulsed electron-electron double resonance (PELDOR) coupled with site-directed spin labeling is a powerful technique for the elucidation of protein or nucleic acid, macromolecular structure and interactions. The intrinsic high sensitivity of electron...
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09-28-2014 08:03 PM
[U. of Ottawa NMR Facility Blog] Measurement of Long Range C H Coupling Constants
Measurement of Long Range C H Coupling Constants
The stereochemistry of compounds is assigned very often with proton - proton NOE's by applying the 2D NOESY technique or the 1D selective gradient NOESY technique. These methods fail, however when the distance between protons is too large to measure an NOE. When faced with this situation, it may be possible to measure long range proton - carbon coupling constants which are able to provide the necessary information. Three-bond carbon - proton couplings follow a Karplus relationship where the magnitude of the coupling constant is related to...
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08-17-2012 10:44 PM
Liquid State DNP for Water Accessibility Measurements on Spin-labeled Membrane Proteins at Physiological Temperatures
Liquid State DNP for Water Accessibility Measurements on Spin-labeled Membrane Proteins at Physiological Temperatures
Publication year: 2012
Source:Journal of Magnetic Resonance</br>
Andrin Doll, Enrica Bordignon, Benesh Joseph, René Tschaggelar, Gunnar Jeschke</br>
We demonstrate the application of continuous wave dynamic nuclear polarization (DNP) at 0.35 Tesla for site-specific water accessibility studies on spin-labeled membrane proteins at concentrations in the 10-100 micromolar range. The DNP effects at such low concentrations are weak and the experimentally...
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[NMR paper] Mapping long-range interactions in alpha-synuclein using spin-label NMR and ensemble
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J Am Chem Soc. 2005 Jan 19;127(2):476-7
Authors: Dedmon MM, Lindorff-Larsen K, Christodoulou J, Vendruscolo M, Dobson CM
The intrinsically disordered protein alpha-synuclein plays a key role in the pathogenesis of Parkinson's disease (PD). We show here that the native state of alpha-synuclein...
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[NMR paper] Measurement of long-range cross-correlation rates using a combination of single- and
Measurement of long-range cross-correlation rates using a combination of single- and multiple-quantum NMR spectroscopy in one experiment.
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J Am Chem Soc. 2002 Apr 17;124(15):4050-7
Authors: Fruh D, Chiarparin E, Pelupessy P, Bodenhausen G
A method is described to determine long-range cross-correlations between the modulations of an anisotropic chemical shift (e.g., of a C' carbonyl carbon in...
A Triarylmethyl Spin Label for Long-Range Distance Measurement at Physiological Temperatures Using T1 Relaxation Enhancement
Linear Mode
