Impact of Post-Translational Succination on Collision-Induced Unfolding of SUMO1

Small Ubiquitin-like Modifier 1 (SUMO1) is a key protein involved in the post-translational modification (PTM) known as SUMOylation, which regulates essential cellular processes such as protein localization, stability, and complex formation. SUMO1 contains a single free cysteine (Cys52) located in an α-helix, which makes it susceptible to succination, another PTM resulting from Michael addition between a thiol group and fumarate, forming 2-(S-succino)cysteine (2SC, Figure 1). Recent studies have shown that succination induces structural alterations in SUMO1 by disrupting its hydrogen-bond network, potentially altering its biological function leading to metabolic alterations. While SUMO1 and its succinated form naturally operate in solution, ion mobility techniques coupled to mass spectrometry (IMS-MS) such as collision-induced unfolding (CIU), probe their structural properties in the gas phase. CIU provides sensitive fingerprint readouts of protein stability, nevertheless these patterns may be difficult to interpret without complementary approaches. In this study, molecular dynamics (MD) simulations across a range of temperatures, used to mimic experimental activation, were performed to gain deeper insight into the structural changes highlighted by CIU profiles. We also applied Monte Carlo simulations to correlate experimental and theoretical CIU50 through the determination of ion temperature in the drift tube following studies of Prell et al.