Development of novel mass spectrometric methods for identifying HOCl-induced modifications to proteins

Laetitia Mouls, Edina Silajdzic, Nicolas Haroune, Corinne M Spickett, Andrew R Pitt

Research output: Contribution to journalArticlepeer-review

Abstract

Protein oxidation is thought to contribute to a number of inflammatory diseases, hence the development of sensitive and specific analytical techniques to detect oxidative PTMs (oxPTMs) in biological samples is highly desirable. Precursor ion scanning for fragment ions of oxidized amino acid residues was investigated as a label-free MS approach to mapping specific oxPTMs in a complex mixture of proteins. Using HOCl-oxidized lysozyme as a model system, it was found that the immonium ions of oxidized tyrosine and tryptophan formed in MS(2) analysis could not be used as diagnostic ions, owing to the occurrence of isobaric fragment ions from unmodified peptides. Using a double quadrupole linear ion trap mass spectrometer, precursor ion scanning was combined with detection of MS(3) fragment ions from the immonium ions and collisionally-activated decomposition peptide sequencing to achieve selectivity for the oxPTMs. For chlorotyrosine, the immonium ion at 170.1 m/z fragmented to yield diagnostic ions at 153.1, 134.1, and 125.1 m/z, and the hydroxytyrosine immonium ion at 152.1 m/z gave diagnostic ions at 135.1 and 107.1 m/z. Selective MS(3) fragment ions were also identified for 2-hydroxytryptophan and 5-hydroxytryptophan. The method was used successfully to map these oxPTMs in a mixture of nine proteins that had been treated with HOCl, thereby demonstrating its potential for application to complex biological samples.
Original languageEnglish
Pages (from-to)1617-1631
Number of pages15
JournalProteomics
Volume9
Issue number6
DOIs
Publication statusPublished - 1 Mar 2009

Keywords

  • 5-hydroxytryptophan
  • amino acid sequence
  • animals
  • hypochlorous acid
  • mass spectrometry
  • molecular sequence data
  • muramidase
  • oxidation-reduction
  • peptides
  • post-translational protein processing
  • proteins
  • protein sequence analysis
  • tyrosine

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