title: Strategies for determination of disulphide bridges in proteins using plasma desorption mass spectrometry.
abstract: Disulphide bridges have been assigned in three different proteins by locating possible disulphide-linked peptides in enzymic digests of the proteins based on their molecular weight determined by plasma desorption mass spectrometry. Different strategies have been employed including in situ reduction of the nitrocellulose-bound peptides and confirmation of peptide identity by methyl esterification reactions or Edman degradation. The latter was needed for identification of glycosylated disulphide-linked peptides. For insulins cleavage between cysteine residues in close proximity was not possible; but a combination of molecular mass information, enzymic cleavage with two different enzymes and sequence analysis including identification of di-phenylthiohydantoin-cystine could ensure an unambiguous assignment of the disulphide bridges.
authors.affiliation: Novo Nordisk a/s, Niels Steensensvej 1, Gentofte, Denmark.
authors.forename: H H
authors.initials: HH
authors.lastname: Sørensen
authors.forename: J
authors.initials: J
authors.lastname: Thomsen
authors.forename: S
authors.initials: S
authors.lastname: Bayne
authors.forename: P
authors.initials: P
authors.lastname: Højrup
authors.forename: P
authors.initials: P
authors.lastname: Roepstorff
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001792:Blood Platelets / Q000737:chemistry; D003545:Cysteine / Q000032:analysis; D004220:Disulfides / Q000032:analysis*; D006023:Glycoproteins / Q000032:analysis; D007328:Insulin / Q000032:analysis; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000032:analysis; D011506:Proteins / Q000032:analysis*; D011994:Recombinant Proteins / Q000032:analysis; D013552:Swine
chemical_list: D004220:Disulfides; D006023:Glycoproteins; D007328:Insulin; D010940:Plant Proteins; D011506:Proteins; D011994:Recombinant Proteins; D003545:Cysteine