Temperature-controlled electrospray ionization tandem mass spectrometry study on protein/small molecule interaction

Abstract

Traditional electrospray ionization tandem mass spectrometry (ESI-MSⁿ) has been a powerful tool in diverse research areas, however, it faces great limitations in the study of protein-small molecule interactions. In this article, the state-of-the-art temperature-controlled electrospray ionization tandem mass spectrometry (TC-ESI-MSⁿ) is applied to investigate interactions between ubiquitin and two flavonol molecules, respectively. The combination of collision-induced dissociation (CID) and MS solution-melting experiments facilitates the understanding of flavonol-protein interactions in a new dimension across varying temperature ranges. While structural changes of proteins disturbed by small molecules are unseen in ESI-MSⁿ, TC-ESI-MSⁿ allows a simultaneous assessment of the stability of the complex in both gas and liquid phases under various temperature conditions, meanwhile investigating the impact on the protein's structure and tracking changes in thermodynamic data, and the characteristics of structural intermediates.