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Despite the growing importance of the mass spectrometry of membrane proteins, it is not known how their transfer from solution into vacuum affects their stability and structure. To address this we have carried out a systematic investigation of ten membrane proteins solubilized in different detergents and used mass spectrometry to gain physicochemical insight into the mechanism of their ionization and desolvation. We show that the chemical properties of the detergents mediate the charge state, both during ionization and detergent removal. Using ion mobility mass spectrometry, we monitor the conformations of membrane proteins and show how the surface charge density dictates the stability of folded states. We conclude that the gas-phase stability of membrane proteins is increased when a greater proportion of their surface is lipophilic and is consequently protected by the physical presence of the micelle., (© 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The study of intact soluble protein assemblies by means of mass spectrometry is providing invaluable contributions to structural biology and biochemistry. A recent breakthrough has enabled similar study of membrane protein complexes, following their release from detergent micelles in the gas phase. Careful optimization of mass spectrometry conditions, particularly with respect to energy regimes, is essential for maintaining compact folded states as detergent is removed. However, many of the saccharide detergents widely employed in structural biology can cause unfolding of membrane proteins in the gas phase. Here, we investigate the potential of charge reduction by introducing three membrane protein complexes from saccharide detergents and show how reducing their overall charge enables generation of compact states, as evidenced by ion mobility mass spectrometry. We find that charge reduction stabilizes the oligomeric state and enhances the stability of lipid-bound complexes. This finding is significant since maintaining native-like membrane proteins enables ligand binding to be assessed from a range of detergents that retain solubility while protecting the overall fold.