Effective novel dissociation methods for intact protein: Heat-assisted nozzle-skimmer collisionally induced dissociation and infrared multiphoton dissociation using a Fourier transform ion cyclotron resonance mass spectrometer equipped with a micrometal e

Heating of a nano-electrospray ionization (nanoESI) source can improve the dissociation efficiency of collisionally induced dissociation (CID) methods, such as nozzle-skimmer CID (NS–CID) and infrared multiphoton dissociation (IRMPD), for large biomolecule fragmentation. A metal nanoESI emitter was used due to its resistance to heating above 250 °C. This novel method for the dissociation of large biomolecular ions is termed “heat-assisted NS–CID” (HANS–CID) or “heat-assisted IRMPD” (HA–IRMPD). Multiple charged nonreduced protein ions (8.6 Da ubiquitin, 14 kDa lysozyme, and 67 kDa bovine serum albumin) were directly dissociated by HANS–CID and HA–IRMPD to effectively yield fragment ions that could be assigned. The fragment ions of ubiquitin by HANS–CID can be analyzed by tandem mass spectrometry (MS/MS) using sustained off-resonance irradiation CID (SORI–CID) and IRMPD. In addition, a native large protein, immunoglobulin G (IgG, 150 kDa), was efficiently dissociated by HA–IRMPD. The product ions that were obtained reflected the domain structure of IgG. However, these product ions of IgG and lysozyme were not dissociated by MS/MS using the same heating energetic methods such as IRMPD and SORI–CID.