The HDX manager was equipped with a Vanguard column (BEH C4, 300 Å, 1.7 m, 2.1 mm x 5 mm; Waters) only. Lipids were trapped on the C4 column and washed with solvent A for 3 minutes at 200 L/min. Subsequently, lipids were eluted by a 3-minute linear gradient from 5 to 95% solvent B (0.23% formic acid in acetonitrile, pH 2.5) at 40 L/min. Eluted lipids were measured in positive ion mode between 50 and 2,000 m/z on the Xevo G2-XS mass spectrometer. The phospholipid trap column was cleaned with 3% NH 4 OH in methanol and re-equilibrated in solvent A during the subsequent wash run. Experiments were performed in the standard two-valve and extended three-valve HDX-MS configuration. Detailed LC settings are provided in Supporting Information (Table and). The extracted ion chromatogram (EIC) of the respective lipid was generated and mass spectra were combined at full width half maximum (FWHM).