Evaluation of Membrane Mimetics for the Native Ion Mobility─Mass Spectrometry Analysis of Membrane Proteins

Abstract

Membrane protein (MP) structures and functions are intricately linked to their surrounding membrane environments and their respective lipid compositions in addition to influences exerted by unique membrane microdomains (i.e., lipid rafts). Owing to the complexity of their native environments, MPs pose significant challenges to structural biology due to their intrinsic hydrophobic nature rendering them incompatible with aqueous conditions commonly deployed during sample preparation. To overcome these solubility issues, MP studies commonly utilize detergent micelles. In recent years, numerous efforts have been directed toward developing membrane mimetics to facilitate successful solubilization and stabilization of MPs. Collision-induced dissociation (CID) can be employed to remove surrounding molecules from MPs during native mass spectrometry (nMS) experiments, and such methods can be used to evaluate previously inaccessible MPs. However, the differential effects of various membrane mimetics on MP structure and stability remain largely unexplored. In this study, we evaluate a range of MPs, including both transmembrane and monotopic variants, solubilized in detergent micelles, mixed lipid bicelles, and nanodisc mimetics. Our findings reveal significant differences in collision induced unfolding (CIU) features, stabilities, collision cross sections (CCS), and RMSD comparisons with values exceeding 26%. These results underscore the importance of considering the choice of membrane mimetics when studying MP structure.