Multi-domain proteins play critical roles in fine-tuning essential processes in cellular signaling and gene regulation. Typically, multiple globular domains that are connected by flexible linkers undergo dynamic re-arrangements upon binding to protein, DNA or RNA ligands. RNA binding proteins (RBPs) represent an important class of multi-domain proteins, which regulate gene expression by recognizing linear or structured RNA sequence motifs. Here, we employ segmental perdeuteration of the three RNA recognition motif (RRM) domains in the RBP TIA-1 using Sortase A-mediated protein ligation. We show that domain-selective perdeuteration combined with contrast-matched small-angle neutron scattering (SANS), SAXS and computational modelling provides valuable information to precisely define relative domain arrangements. The approach is generally applicable to study conformational arrangements of individual domains in multi-domain proteins and changes induced by ligand binding.
[NMR paper] Combining Diffusion NMR and Small-Angle Neutron Scattering Enables Precise Measurements of Polymer Chain Compression in a Crowded Environment.
Combining Diffusion NMR and Small-Angle Neutron Scattering Enables Precise Measurements of Polymer Chain Compression in a Crowded Environment.
Related Articles Combining Diffusion NMR and Small-Angle Neutron Scattering Enables Precise Measurements of Polymer Chain Compression in a Crowded Environment.
Phys Rev Lett. 2017 Mar 03;118(9):097801
Authors: Palit S, He L, Hamilton WA, Yethiraj A, Yethiraj A
Abstract
The effect of particles on the behavior of polymers in solution is important in a number of important phenomena such as...
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[NMR paper] Robust High-Yield Methodologies for (2)H and (2)H/(15)N/(13)C Labeling of Proteins for Structural Investigations Using Neutron Scattering and NMR.
Robust High-Yield Methodologies for (2)H and (2)H/(15)N/(13)C Labeling of Proteins for Structural Investigations Using Neutron Scattering and NMR.
Related Articles Robust High-Yield Methodologies for (2)H and (2)H/(15)N/(13)C Labeling of Proteins for Structural Investigations Using Neutron Scattering and NMR.
Methods Enzymol. 2015;565:3-25
Authors: Duff AP, Wilde KL, Rekas A, Lake V, Holden PJ
Abstract
We have developed a method that has proven highly reliable for the deuteration and triple labeling ((2)H/(15)N/(13)C) of a...
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11-19-2015 05:22 PM
Efficient segmental isotope labeling of multi-domain proteins using Sortase A
Efficient segmental isotope labeling of multi-domain proteins using Sortase A
Abstract
NMR studies of multi-domain protein complexes provide unique insight into their molecular interactions and dynamics in solution. For large proteins domain-selective isotope labeling is desired to reduce signal overlap, but available methods require extensive optimization and often give poor ligation yields. We present an optimized strategy for segmental labeling of multi-domain proteins using the S. aureus transpeptidase Sortase A. Critical improvements compared to...
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08-29-2015 09:18 PM
[NMR paper] Structural Characterization of a Flexible Two-Domain Protein in Solution Using Small Angle X-Ray Scattering and NMR Data.
Structural Characterization of a Flexible Two-Domain Protein in Solution Using Small Angle X-Ray Scattering and NMR Data.
Structural Characterization of a Flexible Two-Domain Protein in Solution Using Small Angle X-Ray Scattering and NMR Data.
Structure. 2014 Nov 6;22(12):1862-1874
Authors: Lemak A, Wu B, Yee A, Houliston S, Lee HW, Gutmanas A, Fang X, Garcia M, Semesi A, Wang YX, Prestegard JH, Arrowsmith CH
Abstract
Multidomain proteins in which individual domains are connected by linkers often possess inherent...
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Structural Characterization of a Flexible Two-Domain Protein in Solution Using Small Angle X-Ray Scattering and NMR Data
Structural Characterization of a Flexible Two-Domain Protein in Solution Using Small Angle X-Ray Scattering and NMR Data
Publication date: Available online 6 November 2014
Source:Structure</br>
Author(s): Alexander Lemak , Bin Wu , Adelinda Yee , Scott Houliston , Hsiau-Wei Lee , Aleksandras Gutmanas , Xianyang Fang , Maite Garcia , Anthony Semesi , Yun-Xing Wang , James*H. Prestegard , Cheryl*H. Arrowsmith</br>
Multidomain proteins in which individual domains are connected by linkers often possess inherent interdomain flexibility that significantly...
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11-07-2014 09:09 AM
[NMR paper] Combining NMR and small angle X-ray and neutron scattering in the structural analysis of a ternary protein-RNA complex.
Combining NMR and small angle X-ray and neutron scattering in the structural analysis of a ternary protein-RNA complex.
Combining NMR and small angle X-ray and neutron scattering in the structural analysis of a ternary protein-RNA complex.
J Biomol NMR. 2013 Mar 3;
Authors: Hennig J, Wang I, Sonntag M, Gabel F, Sattler M
Abstract
Many processes in the regulation of gene expression and signaling involve the formation of protein complexes involving multi-domain proteins. Individual domains that mediate protein-protein and protein-nucleic...
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03-05-2013 03:25 PM
A segmental labeling strategy for unambiguous determination of domainā??domain interactions of large multi-domain proteins
A segmental labeling strategy for unambiguous determination of domainā??domain interactions of large multi-domain proteins
Abstract NMR structural determination of large multi-domain proteins is a challenging task due to significant spectral overlap with a particular difficulty in unambiguous identification of domainā??domain interactions. Segmental labeling is a NMR strategy that allows for isotopically labeling one domain and leaves the other domain unlabeled. This significantly simplifies spectral overlaps and allows for quick identification of domainā??domain interaction. Here, a...
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NMR and Small Angle Scattering-based structural analysis of protein complexes in solu
NMR and Small Angle Scattering-based structural analysis of protein complexes in solution.
Related Articles NMR and Small Angle Scattering-based structural analysis of protein complexes in solution.
J Struct Biol. 2010 Nov 10;
Authors: Madl T, Gabel F, Sattler M
Structural analysis of multi-domain protein complexes is a key challenge in current biology and a prerequisite for understanding the molecular basis of essential cellular processes. The use of solution techniques is important for characterizing the quaternary arrangements and dynamics of...
Segmental, domain-selective perdeuteration and small angle neutron scattering for structural analysis of multi-domain proteins
Linear Mode
