Using covalent modifications to distinguish protein electrospray mechanisms: Charged residue model (CRM) vs. chain ejection model (CEM)

Different mechanisms have been proposed for the formation of gaseous protein ions during electrospray ionization (ESI). In charged residue model (CRM) are produced upon nanodroplet evaporation to dryness. This mechanism is thought dominate in native ESI, where proteins retain compact conformations, with charge states close Rayleigh protein-sized aqueous droplets. Much higher generated from that unfolded solution. The chain ejection (CEM) has ESI under such denaturing conditions. CEM gradually expelled, while mobile H+ equilibrate between droplet and its protruding tail. Providing clear-cut evidence these scenarios remains difficult, because electrosprayed do not usually any features reveal their mechanism. this work we propose stepwise elimination basic sites can serve distinguish CRM CEM. Using cytochrome c as a system, studied had zero 19 Lys blocked by acetylation. (pH 7) same low were observed regardless behavior consistent CRM, governed size, rather than surface chemistry. Denaturing 2) conditions resulted much states. Intriguingly, spectra acquired pH 2 shifted lower when number acetylated was increased. reduction attributed fact lowering compromises ability compete conclusion, illustrate simple covalent modifications help ion via or