title: C-terminal sequencing by mass spectrometry: application to gelatine-derived proline-rich peptides.
abstract: Protonated peptides derived from proline-rich proteins (PRP) are often difficult to sequence by standard collision-induced dissociation (CID) mass spectrometry (MS) due to preferential amide bond cleavage N-terminal to proline. In connection with bovine spongiform encephalopathy regulations, proteolytic products derived from the PRP collagen have been suggested as markers for contamination of animal feedstuffs with processed animal protein (Fernandez Ocaña, M. et al., Analyst 2004, 129, 111-115). Herein, we report the identification of these marker peptides using the strategy of C-terminal sequencing by CID MS from their sodium and lithium adducts. Upon fragmentation a new cationized peptide was produced that is one C-terminal amino acid shorter in length. This dissociation pathway allowed for the facile identification of the C-terminal residue by matrix-assisted laser desorption/ionization tandem time-of-flight mass spectrometry. Each newly formed cationized peptide was further fragmented by up to seven stages of electrospray ionization ion trap MS. Proline-rich C-terminal sequence tags were established which permitted successful database identification of collagen alpha type I proteins.
authors.affiliation: Centre for Chemical and Bioanalytical Sciences, Royal Holloway, University of London, Egham, UK.
authors.forename: Mireia
authors.initials: M
authors.lastname: Fernandez Ocaña
authors.forename: Jackie
authors.initials: J
authors.lastname: Jarvis
authors.forename: Richard
authors.initials: R
authors.lastname: Parker
authors.forename: Peter M
authors.initials: PM
authors.lastname: Bramley
authors.forename: John M
authors.initials: JM
authors.lastname: Halket
authors.forename: Raj K P
authors.initials: RK
authors.lastname: Patel
authors.forename: Hendrik
authors.initials: H
authors.lastname: Neubert
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D015415:Biomarkers; D002417:Cattle; D005780:Gelatin / Q000737:chemistry* / Q000235:genetics; D010446:Peptide Fragments / Q000032:analysis* / Q000235:genetics; D011392:Proline / Q000737:chemistry*; D020539:Sequence Analysis, Protein / Q000379:methods*; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D013552:Swine
chemical_list: D015415:Biomarkers; D010446:Peptide Fragments; D005780:Gelatin; D011392:Proline