Roughhousing with Ions: Surface-Induced Dissociation and Electron Capture Dissociation as Diagnostics of Q-Cyclic...

Native mass spectrometry can characterize a range of biomolecular features pertinent to structural biology, including intact mass, stoichiometry, ligand-bound states, and topology. However, when an instrument's ionization source is tuned to maximize signal intensity or adduct removal, it is possible that the biomolecular complex's tertiary and quaternary structures can be rearranged in a way that no longer reflect its native-like conformation. This could affect downstream ion activation experiments, leading to erroneous conclusions about the native-like structure. One activation strategy is surface-induced dissociation (SID), which generally causes native-like protein complexes to dissociate along the weakest subunit interfaces, revealing critical information about the complex's native-like topology and subunit connectivity. If the quaternary structure has been disturbed, then the SID fingerprint will shift as well. Thus, SID was used to diagnose source-induced quaternary structure rearrangement and help tune an instrument's source and other upstream transmission regions to strike the balance between signal intensity, adduct removal, and conserving the native-like structure. Complementary to SID, electron-capture dissociation (ECD) can also diagnose rearranged quaternary structures and was used after in-source activation to confirm that the subunit interfaces were rearranged, opening the structure to electron capture and subsequent dissociation. These results provide a valuable guide for new practitioners of native mass spectrometry and highlight the importance of using standard protein complexes when tuning new instrument platforms for optimal native mass spectrometry performance.