Hexapeptide libraries for enhanced protein PTM identification and relative abundance profiling in whole human saliva

Sricharan Bandhakavi, Susan K Van Riper, Pierre N. Tawfik, Matthew D. Stone, Tufia Haddad, Nelson L Rhodus, John V Carlis, Timothy J Griffin

Research output: Contribution to journalArticle

34 Citations (Scopus)

Abstract

Dynamic range compression (DRC) by hexapeptide libraries increases MS/MS-based identification of lower-abundance proteins in complex mixtures. However, two unanswered questions impede fully realizing DRCs potential in shotgun proteomics. First, does DRC enhance identification of post-translationally modified proteins? Second, can DRC be incorporated into a workflow enabling relative protein abundance profiling? We sought to answer both questions analyzing human whole saliva. Addressing question one, we coupled DRC with covalent glycopeptide enrichment and MS/MS. With DRC we identified 2 times more N-linked glycoproteins and their glycosylation sites than without DRC, dramatically increasing the known salivary glycoprotein catalog. Addressing question two, we compared differentially stable isotope-labeled saliva samples pooled from healthy and metastatic breast cancer women using a multidimensional peptide fractionation-based workflow, analyzing in parallel one sample portion with DRC and one portion without. Our workflow categorizes proteins with higher absolute abundance, whose relative abundance ratios are altered by DRC, from proteins of lower absolute abundance detected only after DRC. Within each of these salivary protein categories, we identified novel abundance changes putatively associated with breast cancer, demonstrating feasibility and benefits of DRC for relative abundance profiling. Collectively, our results bring us closer to realizing the full potential of DRC for proteomic studies.

Original languageEnglish (US)
Pages (from-to)1052-1061
Number of pages10
JournalJournal of Proteome Research
Volume10
Issue number3
DOIs
StatePublished - Mar 4 2011

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Pulse time modulation
Saliva
Libraries
Workflow
Proteins
Proteomics
Glycoproteins
Salivary Proteins and Peptides
Breast Neoplasms
Glycopeptides
Firearms
Complex Mixtures
Glycosylation
Isotopes
Peptides
Fractionation

Keywords

  • N-linked glycoproteins
  • breast cancer
  • dynamic range compression proteomics
  • proteominer
  • saliva

Cite this

Hexapeptide libraries for enhanced protein PTM identification and relative abundance profiling in whole human saliva. / Bandhakavi, Sricharan; Van Riper, Susan K; Tawfik, Pierre N.; Stone, Matthew D.; Haddad, Tufia; Rhodus, Nelson L; Carlis, John V; Griffin, Timothy J.

In: Journal of Proteome Research, Vol. 10, No. 3, 04.03.2011, p. 1052-1061.

Research output: Contribution to journalArticle

Bandhakavi, Sricharan ; Van Riper, Susan K ; Tawfik, Pierre N. ; Stone, Matthew D. ; Haddad, Tufia ; Rhodus, Nelson L ; Carlis, John V ; Griffin, Timothy J. / Hexapeptide libraries for enhanced protein PTM identification and relative abundance profiling in whole human saliva. In: Journal of Proteome Research. 2011 ; Vol. 10, No. 3. pp. 1052-1061.
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