Development of Novel Free Radical Initiated Peptide Sequencing Reagent: Application to Identification and Characterization of Peptides by Mass Spectrometry

Kaylee Gaspar, Kimberly Fabijanczuk, Tara Otegui, Jose Acosta, Jinshan Gao

Research output: Contribution to journalArticle

3 Scopus citations

Abstract

By incorporating a high proton affinity moiety to the charge localized free radical-initiated peptide sequencing (CL-FRIPS) reagent, FRIPS-MS technique has extended the applicability to hydrophobic peptides and peptides without basic amino acid residues (lysine, arginine, and histidine). Herein, the CL-FRIPS reagent has three moieties: (1) pyridine acting as the basic site to locate the proton, (2) 2,2,6,6-tetramethylpiperidine-1-oxyl (TEMPO, a stable free radical) acting as the free radical precursor to generate the nascent free radical in the gas phase, and (3) N-hydroxysuccinimide (NHS) activated carboxylic acid acting as the coupling site to derivatize the N-terminus of peptides. The CL-FRIPS reagent allows for the characterization of peptides by generating sequencing ions, enzymatic cleavage-like radical-induced side chain losses, and the loss of TEMPO simultaneously via one-step collisional activation. Further collisional activation of enzymatic cleavage-like radical-induced side chain loss ions provides more information for the structure determination of peptides. The application of CL-FRIPS reagent to characterize peptides is proved by employing bovine insulin as the model peptide. Both scaffold structure of bovine insulin and sequencing information of each chain are achieved. [Figure not available: see fulltext.].

Original languageEnglish
Pages (from-to)548-556
Number of pages9
JournalJournal of the American Society for Mass Spectrometry
Volume30
Issue number3
DOIs
StatePublished - 15 Mar 2019

Keywords

  • Charge localize
  • Free radical
  • Hydrophobic peptides
  • Insulin
  • Peptide sequencing
  • Peptides without basic amino acid residues

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