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Updated: Jul 26, 2025

Analyzing Protein Architectures and Protein-Ligand Complexes by Integrative Structural Mass Spectrometry
Published on: October 15, 2018
Tracking the mechanism of covalent molecular glue stabilization using native mass spectrometry
Carlo J A Verhoef1, Danielle F Kay2, Lars van Dijck1
1Department of Biomedical Engineering and Institute for Complex Molecular Systems, Eindhoven University of Technology Eindhoven 5600 MB The Netherlands p.cossar@tue.nl.
Abstract:
Molecular glues are powerful tools for the control of protein-protein interactions. Yet, the mechanisms underlying multi-component protein complex formation remain poorly understood. Native mass spectrometry (MS) detects multiple protein species simultaneously, providing an entry to elucidate these mechanisms. Here, for the first time, covalent molecular glue stabilization was kinetically investigated by combining native MS with biophysical and structural techniques. This approach elucidated the stoichiometry of a multi-component protein-ligand complex, the assembly order, and the contributions of covalent versus non-covalent binding events that govern molecular glue activity. Aldehyde-based molecular glue activity is initially regulated by cooperative non-covalent binding, followed by slow covalent ligation, further enhancing stabilization. This study provides a framework to investigate the mechanisms of covalent small molecule ligation and informs (covalent) molecular glue development.
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