In the last three decades, the scope of solid-state NMR has expanded to exploring complex biomolecules, from large protein assemblies to intact cells at atomic-level resolution. This diversity in macromolecules frequently features highly flexible components whose insoluble environment precludes the use of solution NMR to study their structure and interactions. While High-resolution Magic-Angle Spinning (HR-MAS) probes offer the capacity for gradient-based 1H-detected spectroscopy in solids, such probes are not commonly used for routine MAS NMR experiments. As a result, most exploration of the flexible regime entails either 13C-detected experiments, the use of partially perdeuterated systems, or ultra-fast MAS. Here we explore proton-detected pulse schemes probing through-bond 13Cā??13C networks to study mobile protein sidechains as well as polysaccharides in a broadband manner. We demonstrate the use of such schemes to study a mixture of microtubule-associated protein (MAP) tau and human microtubules (MTs), and the cell wall of the fungus Schizophyllum commune using 2D and 3D spectroscopy, to show its viability for obtaining unambiguous correlations using standard fast-spinning MAS probes at high and ultra-high magnetic fields.
[NMR paper] 1H-detected characterization of carbon-carbon networks in highly flexible protonated biomolecules using MAS NMR
1H-detected characterization of carbon-carbon networks in highly flexible protonated biomolecules using MAS NMR
In the last three decades, the scope of solid-state NMR has expanded to exploring complex biomolecules, from large protein assemblies to intact cells at atomic-level resolution. This diversity in macromolecules frequently features highly flexible components whose insoluble environment precludes the use of solution NMR to study their structure and interactions. While High-resolution Magic-Angle Spinning (HR-MAS) probes offer the capacity for gradient-based ¹H-detected spectroscopy...
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06-08-2023 09:28 PM
[NMR paper] Interdomain Dynamics via Paramagnetic NMR on the Highly Flexible Complex Calmodulin/Munc13-1
Interdomain Dynamics via Paramagnetic NMR on the Highly Flexible Complex Calmodulin/Munc13-1
Paramagnetic NMR constraints are very useful to study protein interdomain motion, but their interpretation is not always straightforward. On the example of the particularly flexible complex Calmodulin/Munc13-1, we present a new approach to characterize this motion with pseudocontact shifts and residual dipolar couplings. Using molecular mechanics, we sampled the conformational space of the complex and used a genetic algorithm to find ensembles that are in agreement with the data. We used the...
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09-11-2022 10:03 PM
NMR mapping of the highly flexible regions of 13 C/ 15 N-labeled antibody TTAC-0001-Fab
NMR mapping of the highly flexible regions of 13 C/ 15 N-labeled antibody TTAC-0001-Fab
Abstract
Monoclonal antibody (mAb) drugs are clinically important for the treatment of various diseases. TTAC-0001 is under development as a new anti-cancer antibody drug targeting VEGFR-2. As the less severe toxicity of TTAC-0001 compared to Bevacizumab, likely due to the decreased in vivo half-life, seems to be related to its structural flexibility, it is important to map the exact flexible regions. Although the 13C/15N-labeled protein is required for NMR...
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05-16-2020 02:10 AM
Deleterious effects of carbonā??carbon dipolar coupling on RNA NMR dynamics
Deleterious effects of carbonā??carbon dipolar coupling on RNA NMR dynamics
Abstract
Many regulatory RNAs undergo dynamic exchanges that are crucial for their biological functions and NMR spectroscopy is a versatile tool for monitoring dynamic motions of biomolecules. Meaningful information on biomolecular dynamics requires an accurate measurement of relaxation parameters such as longitudinal (R1) rates, transverse (R2) rates and heteronuclear Overhauser effect (hNOE). However, earlier studies have shown that the large 13Cā??13C interactions...
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05-03-2020 02:46 PM
Asynchronous through-bond homonuclear isotropic mixing: application to carbonā??carbon transfer in perdeuterated proteins under MAS
Asynchronous through-bond homonuclear isotropic mixing: application to carbonā??carbon transfer in perdeuterated proteins under MAS
Abstract
Multiple-bond carbonā??carbon homonuclear mixing is a hurdle in extensively deuterated proteins and under fast MAS due to the absence of an effective proton dipolar-coupling network. Such conditions are now commonly employed in solid-state NMR spectroscopy. Here, we introduce an isotropic homonuclear 13Cā??13C through-bond mixing sequence, MOCCA, for the solid state. Even though applied under MAS, this scheme...
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08-29-2015 09:18 PM
ā??CON-CONā?? assignment strategy for highly flexible intrinsically disordered proteins
ā??CON-CONā?? assignment strategy for highly flexible intrinsically disordered proteins
Abstract
Intrinsically disordered proteins (IDPs) are a class of highly flexible proteins whose characterization by NMR spectroscopy is complicated by severe spectral overlaps. The development of experiments designed to facilitate the sequence-specific assignment procedure is thus very important to improve the tools for the characterization of IDPs and thus to be able to focus on IDPs of increasing size and complexity. Here, we present and describe the...
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10-21-2014 11:31 PM
[NMR paper] Global folds of highly deuterated, methyl-protonated proteins by multidimensional NMR
Global folds of highly deuterated, methyl-protonated proteins by multidimensional NMR.
http://www.ncbi.nlm.nih.gov/corehtml/query/egifs/http:--pubs.acs.org-images-acspubs.jpg Related Articles Global folds of highly deuterated, methyl-protonated proteins by multidimensional NMR.
Biochemistry. 1997 Feb 11;36(6):1389-401
Authors: Gardner KH, Rosen MK, Kay LE
The development of 15N, 13C, 2H multidimensional NMR spectroscopy has facilitated the assignment of backbone and side chain resonances of proteins and protein complexes with molecular masses...
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08-22-2010 03:31 PM
[NMR paper] Global folds of highly deuterated, methyl-protonated proteins by multidimensional NMR
Global folds of highly deuterated, methyl-protonated proteins by multidimensional NMR.
http://www.ncbi.nlm.nih.gov/corehtml/query/egifs/http:--pubs.acs.org-images-acspubs.jpg Related Articles Global folds of highly deuterated, methyl-protonated proteins by multidimensional NMR.
Biochemistry. 1997 Feb 11;36(6):1389-401
Authors: Gardner KH, Rosen MK, Kay LE
The development of 15N, 13C, 2H multidimensional NMR spectroscopy has facilitated the assignment of backbone and side chain resonances of proteins and protein complexes with molecular masses...