Abstract
Intrinsically disordered proteins (IDPs) play important roles in the regulation of cellular function and in disease, and thus they represent an important group of therapeutic targets. Yet, members of this “disorderome” have not yet been successfully targeted by drugs, primarily because traditional design principles cannot be applied to their highly dynamic, heterogeneous structural states. Binders developed against IDPs so far suffer from very weak binding and inability to act in a cellular context. Here, we describe a possible generic method for the targeting of IDPs via covalent modification, which could entail specific and strong binding and inhibitory potential, making such “warheads” reasonable starting points of drug-development efforts. We demonstrate this principle by addressing the cysteine-specific covalent modification of calpastatin, the IDP inhibitor of the calcium-dependent cysteine protease calpain. We describe the protocol for monitoring the covalent modification of the inhibitor, measuring the Ki of its inhibition and comparing it to the Kd of its interaction with the enzyme. Our premise is that the underlying principles can be easily adapted to screen for molecules targeting other, disease-related, IDPs in the future.
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Acknowledgements
This work was supported by the Odysseus grant G.0029.12 and the postdoctoral fellowship #1218713 from Research Foundation Flanders (FWO), the H2020 MSCA ITN FRAGNET (project 6758993) grant and K124670 and PD124598 grants from the Hungarian Scientific Research Fund (OTKA). We thank Wim Versées, Joris Messens, and Maria Tossounian for helpful discussions.
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Nguyen, H.H. et al. (2020). Targeting an Intrinsically Disordered Protein by Covalent Modification. In: Kragelund, B.B., Skriver, K. (eds) Intrinsically Disordered Proteins. Methods in Molecular Biology, vol 2141. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0524-0_43
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DOI: https://doi.org/10.1007/978-1-0716-0524-0_43
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