title: The identification of disulfides in ricin D using proteolytic cleavage followed by negative-ion nano-electrospray ionization mass spectrometry of the peptide fragments.
abstract: BACKGROUND
  To use negative-ion nano-electrospray ionization mass spectrometry of peptides from the tryptic digest of ricin D, to provide sequence information; in particular, to identify disulfide position and connectivity.
  
  
  METHODS
  Negative-ion fragmentations of peptides from the tryptic digest of ricin D was studied using a Waters QTOF2 mass spectrometer operating in MS and MS(2) modes.
  
  
  RESULTS
  Twenty-three peptides were obtained following high-performance liquid chromatography and studied by negative-ion mass spectrometry covering 73% of the amino-acid residues of ricin D. Five disulfide-containing peptides were identified, three intermolecular and two intramolecular disulfide-containing peptides. The [M-H](-) anions of the intermolecular disulfides undergo facile cleavage of the disulfide units to produce fragment peptides. In negative-ion collision-induced dissociation (CID) these source-formed anions undergo backbone cleavages, which provide sequencing information. The two intramolecular disulfides were converted proteolytically into intermolecular disulfides, which were identified as outlined above.
  
  
  CONCLUSIONS
  The positions of the five disulfide groups in ricin D may be determined by characteristic negative-ion cleavage of the disulfide groups, while sequence information may be determined using the standard negative-ion backbone cleavages of the resulting cleaved peptides. Negative-ion mass spectrometry can also be used to provide partial sequencing information for other peptides (i.e. those not containing Cys) using the standard negative-ion backbone cleavages of these peptides.
authors: Department of Chemistry, The University of Adelaide, South Australia, Australia, 5005.
authors: T T Nha
authors: TT
authors: Tran
authors: Craig S
authors: CS
authors: Brinkworth
authors: John H
authors: JH
authors: Bowie
mesh_terms: D000595:Amino Acid Sequence; D004220:Disulfides / Q000032:analysis* / Q000737:chemistry; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000032:analysis* / Q000737:chemistry* / Q000378:metabolism; D012276:Ricin / Q000032:analysis* / Q000737:chemistry* / Q000378:metabolism; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D014357:Trypsin
chemical_list: D004220:Disulfides; D010446:Peptide Fragments; D012276:Ricin; D014357:Trypsin