Structure and dynamics of an intrinsically disordered protein region that partially folds upon binding by chemical-exchange NMR

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Structure and dynamics of an intrinsically disordered protein region that partially folds upon binding by chemical-exchange NMR, Journal of the American Chemical SocietyVolume 139, p. 12219 - 12227, 2017

 

Intrinsically disordered proteins (IDPs) and regions (IDRs) are ubiquitous in the proteome. The flexibility of IDPs allows them to engage in many interactions with other biomolecules. These interactions often lead to folding of a local sequence motif but most parts of a disordered protein usually retain some disorder. In extreme cases, an entire IDP assembles in a fuzzy complex. The presence of both ordered and disordered segments makes the characterization of conformational ensembles of the bound states of IDPs challenging.6 A widely applicable method to determine the conformational properties of the interacting regions of complexes involving long IDRs in solution is still missing.

 

 

Here, we introduce a method for the structural determination of complexes consisting of disordered protein regions that fold in part upon binding to a partner protein. This approach combines chemical-exchange NMR, structural determination of the interacting region of the disordered protein, and docking. Importantly, the method does not require the observation of the bound form of the IDP and provides structural insight on both fully and partially ordered regions in the bound IDP. We apply this method to a complex between a 96-residue disordered region of Artemis and the DBD of Ligase IV. We show that our approach correctly reproduces the crystal structure of the folded core of the complex and provides additional information on regions within the IDP that contribute to the thermodynamic affinity but retain some disorder. The method is not affected by edge effects when short peptides are used in a more traditional approach. From our NMR and ITC-based investigations, we identify additional residues at the Cterminal end of Artemis (496-501) that contribute significantly to the binding to Ligase IV. This NMR-based approach to the structural characterization of complexes of IDPs is robust and is expected to be applicable to a wide range of systems.

 

Pour plus d'information, consultez l'actualité vulgarisée rédigée en collaboration avec le CNRS : Comment les protéines desordonnées enlacent leur partenaire ?

 

Résumé: 

Journal of the American Chemical Society, Volume 139, p. 12219 - 12227, 2017

 

Many intrinsically disordered proteins (IDPs) and protein regions (IDRs) engage in transient, yet specific, interactions with a variety of protein partners. Often, if not always, interactions with a protein partner lead to partial folding of the IDR. Characterizing the conformational space of such complexes is challenging: in solution-state NMR, signals of the IDR in the interacting region become broad, weak and often invisible; while X-ray crystallography only provides information on fully ordered regions. There is thus a need for a simple method to characterize both fully and partially ordered regions in the bound state of IDPs. Here, we introduce an approach based on monitoring chemical exchange by NMR to investigate the state of an IDR that folds upon binding through the observation of the free state of the protein. Structural constraints for the bound state are obtained from chemical shifts and site-specific dynamics of the bound state are characterized by relaxation rates. The conformation of the interacting part of the IDR was determined and subsequently docked onto the structure of the folded partner. We apply the method to investigate the interaction between the disordered C-terminal region of Artemis and the DNA binding domain of Ligase IV. We show that we can accurately reproduce the structure of the core of the complex determined by X-ray crystallography and identify a broader interface. The method is widely applicable to the biophysical investigation of complexes of disordered proteins and folded proteins.

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Structure and dynamics of an intrinsically disordered protein region that partially folds upon binding by chemical-exchange NMR

 

Cyril Charlier, Guillaume Bouvignies, Philippe Pelupessy, Astrid Walrant, Rodrigue Marquant, Mikhail Kozlov, Pablo De Ioannes, Nicolas Bolik-Coulon, Sandrine Sagan, Patricia Cortes, Aneel K. Aggarwal, Ludovic Carlier, and Fabien Ferrage

 

Journal of the American Chemical Society, Volume 139, p. 12219 - 12227, 2017

 

DOI :10.1021/jacs.7b05823