title: Electron-capture dissociation and ion mobility mass spectrometry for characterization of the hemoglobin protein assembly.
abstract: Native spray has the potential to probe biophysical properties of protein assemblies. Here we report an investigation using both ECD top-down sequencing with an FTICR mass spectrometer and ion mobility (IM) measurements on a Q-TOF to investigate the collisionally induced unfolding of a native-like heterogeneous tetrameric assembly, human hemoglobin (hHb), in the gas phase. To our knowledge, this is the first report combining ECD and ion-mobility data on the same target protein assembly to delineate the effects of collisional activation on both assembly size and the extent and location of fragmentation. Although the collision-induced unfolding of the hemoglobin assembly is clearly seen by both IMMS and ECD, the latter delineates the regions that increasingly unfold as the collision energy is increased. The results are consistent with previous outcomes for homogeneous protein assemblies and reinforce our interpretation that activation opens the structure of the protein assembly from the flexible regions to make available ECD fragmentation, without dissociating the component proteins.
authors.affiliation: Department of Chemistry, Washington University in St. Louis, St. Louis, Missouri, 63130.
authors.forename: Weidong
authors.initials: W
authors.lastname: Cui
authors.affiliation: Department of Chemistry, Washington University in St. Louis, St. Louis, Missouri, 63130.
authors.forename: Hao
authors.initials: H
authors.lastname: Zhang
authors.affiliation: Department of Chemistry, Washington University in St. Louis, St. Louis, Missouri, 63130.|Department of Biology, Washington University in St. Louis, St. Louis, Missouri, 63130.
authors.forename: Robert E
authors.initials: RE
authors.lastname: Blankenship
authors.affiliation: Department of Chemistry, Washington University in St. Louis, St. Louis, Missouri, 63130.
authors.forename: Michael L
authors.initials: ML
authors.lastname: Gross
mesh_terms: D000595:Amino Acid Sequence; D018360:Crystallography, X-Ray; D006454:Hemoglobins / Q000737:chemistry*; D006801:Humans; D008958:Models, Molecular; D055503:Protein Multimerization; D017433:Protein Structure, Secondary; D021241:Spectrometry, Mass, Electrospray Ionization
chemical_list: D006454:Hemoglobins
keywords: B factor; electron capture dissociation; hemoglobin; ion mobility; mass spectrometry; protein assembly