title: On-line electrochemical reduction of disulfide bonds: improved FTICR-CID and -ETD coverage of oxytocin and hepcidin.
abstract: Particularly in the field of middle- and top-down peptide and protein analysis, disulfide bridges can severely hinder fragmentation and thus impede sequence analysis (coverage). Here we present an on-line/electrochemistry/ESI-FTICR-MS approach, which was applied to the analysis of the primary structure of oxytocin, containing one disulfide bridge, and of hepcidin, containing four disulfide bridges. The presented workflow provided up to 80% (on-line) conversion of disulfide bonds in both peptides. With minimal sample preparation, such reduction resulted in a higher number of peptide backbone cleavages upon CID or ETD fragmentation, and thus yielded improved sequence coverage. The cycle times, including electrode recovery, were rapid and, therefore, might very well be coupled with liquid chromatography for protein or peptide separation, which has great potential for high-throughput analysis.
authors.affiliation: Leiden University Medical Center (LUMC), Center for Proteomics and Metabolomics, 2300 RC, Leiden, The Netherlands.
authors.forename: Simone
authors.initials: S
authors.lastname: Nicolardi
authors.forename: Martin
authors.initials: M
authors.lastname: Giera
authors.forename: Pieter
authors.initials: P
authors.lastname: Kooijman
authors.forename: Agnieszka
authors.initials: A
authors.lastname: Kraj
authors.forename: Jean-Pierre
authors.initials: JP
authors.lastname: Chervet
authors.forename: André M
authors.initials: AM
authors.lastname: Deelder
authors.forename: Yuri E M
authors.initials: YE
authors.lastname: van der Burgt
mesh_terms: D000595:Amino Acid Sequence; D004220:Disulfides / Q000737:chemistry*; D055664:Electrochemical Techniques / Q000295:instrumentation*; D004867:Equipment Design; D064451:Hepcidins / Q000737:chemistry*; D013058:Mass Spectrometry / Q000295:instrumentation*; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D010121:Oxytocin / Q000737:chemistry*
chemical_list: D004220:Disulfides; D064451:Hepcidins; D010121:Oxytocin
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