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 acid interactions are typically connected by flexible linkers, which contribute to conformational dynamics and enable the formation of complexes with distinct binding partners. Solution techniques are therefore required for structural analysis and to characterize potential conformational dynamics. Nuclear magnetic resonance spectroscopy (NMR) provides such information but often only sparse data are obtained with increasing molecular weight of the complexes. It is therefore beneficial to combine NMR data with additional structural restraints from complementary solution techniques. Small angle X-ray/neutron scattering (SAXS/SANS) data can be efficiently combined with NMR-derived information, either for validation or by providing additional restraints for structural analysis. Here, we show that the combination of SAXS and SANS data can help to refine structural models obtained from data-driven docking using HADDOCK based on sparse NMR data. The approach is demonstrated with the ternary protein-protein-RNA complex involving two RNA recognition motif (RRM) domains of Sex-lethal, the N-terminal cold shock domain of Upstream-to-N-Ras, and msl-2 mRNA. Based on chemical shift perturbations we have mapped protein-protein and protein-RNA interfaces and complemented this NMR-derived information with SAXS data, as well as SANS measurements on subunit-selectively deuterated samples of the ternary complex. Our results show that, while the use of SAXS data is beneficial, the additional combination with contrast variation in SANS data resolves remaining ambiguities and improves the docking based on chemical shift perturbations of the ternary protein-RNA complex.
PMID: 23456097 [PubMed - as supplied by publisher]
Small Angle X-Ray Scattering and the Determination of Protein Shape - Azom.com
Small Angle X-Ray Scattering and the Determination of Protein Shape - Azom.com
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Small Angle X-Ray Scattering and the Determination of Protein Shape
Azom.com
The low resolution 3D structure obtained by ab-initio methods is an important additional data to the high resolution structures obtained by protein crystallography and NMR. This article discusses the application of SAXSess mc system from Anton Paar to ...
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Contribution from the Laboratory of Chemical Physics, NIDDK, National Institutes of Health, Bethesda, Maryland 20892-0520, Department of Biochemistry, The Ohio State University, Columbus, Ohio 43210, and Department of Chemistry, University of Utah, Salt Lake City, Utah 84112-0850
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