Effects of single amino acid substitution on the collision-induced dissociation of intact protein ions: Turkey ovomucoid third domain

J Proteome Res. Sep-Oct 2004;3(5):1033-41. doi: 10.1021/pr049910w.


Expanded understanding of the factors that direct polypeptide ion fragmentation can lead to improved specificity in the use of tandem mass spectrometry for the identification and characterization of proteins. Like the fragmentation of peptide cations, the dissociation of whole protein cations shows several preferred cleavages, the likelihood for which is parent ion charge dependent. While such cleavages are often observed, they are far from universally observed, despite the presence of the residues known to promote them. Furthermore, cleavages at residues not noted to be common in a variety of proteins can be dominant for a particular protein or protein ion charge state. Motivated by the ability to study a small protein, turkey ovomucoid third domain, for which a variety of single amino acid variants are available, the effects of changing the identity of one amino acid in the protein sequence on its dissociation behavior were examined. In particular, changes in amino acids associated with C-terminal aspartic acid cleavage and N-terminal proline cleavage were emphasized. Consistent with previous studies, the product ion spectra were found to be dependent upon the parent ion charge state. Furthermore, the fraction of possible C-terminal aspartic acid cleavages observed to occur for this protein was significantly larger than the fraction of possible N-terminal proline cleavages. In fact, very little N-terminal proline cleavage was noted for the wild-type protein despite the presence of three proline residues in the protein. The addition/removal of proline and aspartic acids was studied along with changes in selected residues adjacent to proline residues. Evidence for inhibition of proline cleavage by the presence of nearby basic residues was noted, particularly if the basic residue was likely to be protonated.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Amino Acid Substitution*
  • Animals
  • Arginine / chemistry
  • Aspartic Acid / chemistry
  • Histidine / chemistry
  • Lysine / chemistry
  • Mass Spectrometry*
  • Molecular Sequence Data
  • Ovomucin / chemistry*
  • Ovomucin / genetics
  • Oxidation-Reduction
  • Peptide Fragments / chemistry
  • Peptide Fragments / genetics
  • Proline / chemistry
  • Recombinant Proteins / chemistry
  • Turkeys


  • Peptide Fragments
  • Recombinant Proteins
  • Aspartic Acid
  • Ovomucin
  • Histidine
  • Arginine
  • Proline
  • Lysine