Moving away from the reference genome: Evaluating a peptide sequencing tagging approach for single amino acid polymorphism identifications in the genus populus

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Abstract

The genetic diversity across natural populations of the model organism, Populus, is extensive, containing a single nucleotide polymorphism roughly every 200 base pairs. When deviations from the reference genome occur in coding regions, they can impact protein sequences. Rather than relying on a static reference database to profile protein expression, we employed a peptide sequence tagging (PST) approach capable of decoding the plasticity of the Populus proteome. Using shotgun proteomics data from two genotypes of P. trichocarpa, a tag-based approach enabled the detection of 6653 unexpected sequence variants. Through manual validation, our study investigated how the most abundant chemical modification (methionine oxidation) could masquerade as a sequence variant (Ala→Ser) when few site-determining ions existed. In fact, precise localization of an oxidation site for peptides with more than one potential placement was indeterminate for 70% of the MS/MS spectra. We demonstrate that additional fragment ions made available by high energy collisional dissociation enhances the robustness of the peptide sequence tagging approach (81% of oxidation events could be exclusively localized to a methionine). We are confident that augmenting fragmentation processes for a PST approach will further improve the identification of single amino acid polymorphism in Populus and potentially other species as well.

Original languageEnglish
Pages (from-to)3642-3651
Number of pages10
JournalJournal of Proteome Research
Volume12
Issue number8
DOIs
StatePublished - Aug 2 2013

Keywords

  • Populus
  • high energy collisional dissociation
  • mass spectrometry
  • peptide sequence tagging
  • plant proteomics
  • single amino acid polymorphisms

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