title: A proteomics grade electron transfer dissociation-enabled hybrid linear ion trap-orbitrap mass spectrometer.
abstract: Here we detail the modification of a quadrupole linear ion trap-orbitrap hybrid (QLT-orbitrap) mass spectrometer to accommodate a negative chemical ionization (NCI) source. The NCI source is used to produce fluoranthene radical anions for imparting electron transfer dissociation (ETD). The anion beam is stable, robust, and intense so that a sufficient amount of reagents can be injected into the QLT in only 4-8 ms. Following ion/ion reaction in the QLT, ETD product ions are mass-to-charge (m/z) analyzed in either the QLT (for speed and sensitivity) or the orbitrap (for mass resolution and accuracy). Here we describe the physical layout of this device, parametric optimization of anion transport, an evaluation of relevant ETD figures of merit, and the application of this instrument to protein sequence analysis. Described proteomic applications include complex peptide mixture analysis, post-translational modification (PTM) site identification, isotope-encoded quantitation, large peptide characterization, and intact protein analysis. From these experiments, we conclude the ETD-enabled orbitrap will provide the proteomic field with several new opportunities and represents an advance in protein sequence analysis technologies.
authors.affiliation: Department of Chemistry and Biomolecular Chemistry, University of Wisconsin, Madison, WI 53706, USA.
authors.forename: Graeme C
authors.initials: GC
authors.lastname: McAlister
authors.forename: W Travis
authors.initials: WT
authors.lastname: Berggren
authors.forename: Jens
authors.initials: J
authors.lastname: Griep-Raming
authors.forename: Stevan
authors.initials: S
authors.lastname: Horning
authors.forename: Alexander
authors.initials: A
authors.lastname: Makarov
authors.forename: Doug
authors.initials: D
authors.lastname: Phanstiel
authors.forename: George
authors.initials: G
authors.lastname: Stafford
authors.forename: Danielle L
authors.initials: DL
authors.lastname: Swaney
authors.forename: John E P
authors.initials: JE
authors.lastname: Syka
authors.forename: Vlad
authors.initials: V
authors.lastname: Zabrouskov
authors.forename: Joshua J
authors.initials: JJ
authors.lastname: Coon
mesh_terms: D000595:Amino Acid Sequence; D002247:Carbon Isotopes; D002460:Cell Line; D045744:Cell Line, Tumor; D045424:Complex Mixtures / Q000032:analysis; D004583:Electrons; D053595:Embryonic Stem Cells / Q000737:chemistry; D004867:Equipment Design; D005449:Fluorenes / Q000737:chemistry; D006657:Histones / Q000032:analysis; D006801:Humans; D007477:Ions; D007553:Isotope Labeling; D008969:Molecular Sequence Data; D010748:Phosphopeptides / Q000032:analysis; D040901:Proteomics / Q000295:instrumentation*; D029701:Saccharomyces cerevisiae Proteins / Q000032:analysis; D053719:Tandem Mass Spectrometry / Q000295:instrumentation*
chemical_list: D002247:Carbon Isotopes; D045424:Complex Mixtures; D005449:Fluorenes; D006657:Histones; D007477:Ions; D010748:Phosphopeptides; D029701:Saccharomyces cerevisiae Proteins; C007738:fluoranthene