title: Ion mobility-mass spectrometry (IM-MS) for top-down proteomics: increased dynamic range affords increased sequence coverage.
abstract: A general approach that combines mass spectrometry (MS), collision-induced dissociation (CID), ion mobility (IM), and MS for top-down proteomics is described, denoted as MS-CID-IM-MS. Using this approach, CID product ions are dispersed in two dimensions, specifically size-to-charge (IM) and mass-to-charge (MS), and the resulting 2D data display greatly facilitates peptide/protein mass mapping, amino acid sequence analysis, and determination of site-specific protein modifications. Also, this approach alleviates some of the inherent limitations of top-down proteomics, viz. the limitations in dynamic range for fragment ion abundances owing to the number of fragmentation channels available to large ionic systems as well as the resulting spectral congestion. For large peptides such as melittin (2845 Da), CID of the [M + 3H](3+), [M + 4H](4+), and [M + 5H](5+) ions yields amino acid sequence coverage of 42.3%, 38.5%, and 7.7%, respectively, whereas the hybrid MS-CID-IM-MS approach yields amino acid sequence coverages of 84.6%, 65.4%, and 69.2%, respectively. For large biomolecules such as ubiquitin (8565 Da), the amino acid sequence coverage increases from 39% to 76%. The MS-CID-IM-MS top-down approach allows for greater depth of information by allowing the assignment and study of internal fragment ions. Lastly, analysis of the methyl esterification of ubiquitin and single point mutation of human iron sulfur cluster U (HISCU, 14.3 kDa) demonstrates the ability of MS-CID-IM-MS to rapidly identify the presence and sites of modifications.
authors.affiliation: Laboratory for Biological Mass Spectrometry, Department of Chemistry, Texas A&M University, College Station, Texas, USA.
authors.forename: Nathanael F
authors.initials: NF
authors.lastname: Zinnel
authors.forename: Pei-Jing
authors.initials: PJ
authors.lastname: Pai
authors.forename: David H
authors.initials: DH
authors.lastname: Russell
mesh_terms: D000595:Amino Acid Sequence; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods*; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry; D011506:Proteins / Q000737:chemistry; D040901:Proteomics / Q000379:methods*; D013997:Time Factors
chemical_list: D010446:Peptide Fragments; D011506:Proteins