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J Am Chem Soc. 2017 Sep 6;139(35):12165-12174. doi: 10.1021/jacs.7b03875. Epub 2017 Aug 25.

Characterization of Protein-Protein Interfaces in Large Complexes by Solid-State NMR Solvent Paramagnetic Relaxation Enhancements.

Author information

1
Department of Chemistry, University of Warwick , Gibbet Hill Road, Coventry CV4 7AL, U.K.
2
Center for Integrated Protein Science, Department of Chemistry, Munich Technische Universität München , Lichtenbergstrasse 4, 85748 Garching, Germany.
3
Institute of Structural Biology, Helmholtz Zentrum München , Ingolstädter Landstrasse 1, 85764 Neuherberg, Germany.
4
Department of Physics, University of Warwick , Gibbet Hill Road, Coventry CV4 7AL, U.K.
5
Institute of Health Technologies, Tallinn University of Technology , Akadeemia tee 15a, 19086 Tallinn, Estonia.
6
Institute of Molecular Biology and Biochemistry, Center of Molecular Medicine, Medical University of Graz , Harrachgasse 21, 8010 Graz, Austria.

Abstract

Solid-state NMR is becoming a viable alternative for obtaining information about structures and dynamics of large biomolecular complexes, including ones that are not accessible to other high-resolution biophysical techniques. In this context, methods for probing protein-protein interfaces at atomic resolution are highly desirable. Solvent paramagnetic relaxation enhancements (sPREs) proved to be a powerful method for probing protein-protein interfaces in large complexes in solution but have not been employed toward this goal in the solid state. We demonstrate that 1H and 15N relaxation-based sPREs provide a powerful tool for characterizing intermolecular interactions in large assemblies in the solid state. We present approaches for measuring sPREs in practically the entire range of magic angle spinning frequencies used for biomolecular studies and discuss their benefits and limitations. We validate the approach on crystalline GB1, with our experimental results in good agreement with theoretical predictions. Finally, we use sPREs to characterize protein-protein interfaces in the GB1 complex with immunoglobulin G (IgG). Our results suggest the potential existence of an additional binding site and provide new insights into GB1:IgG complex structure that amend and revise the current model available from studies with IgG fragments. We demonstrate sPREs as a practical, widely applicable, robust, and very sensitive technique for determining intermolecular interaction interfaces in large biomolecular complexes in the solid state.

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