Large-scale phosphoproteomics analysis of whole saliva reveals a distinct phosphorylation pattern

Matthew D. Stone, Xiaobing Chen, Thomas McGowan, Sricharan Bandhakavi, Bin Cheng, Nelson L Rhodus, Timothy J Griffin

Research output: Contribution to journalArticle

32 Citations (Scopus)

Abstract

In-depth knowledge of bodily fluid phosphoproteomes, such as whole saliva, is limited. To better understand the whole saliva phosphoproteome, we generated a large-scale catalog of phosphorylated proteins. To circumvent the wide dynamic range of phosphoprotein abundance in whole saliva, we combined dynamic range compression using hexapeptide beads, strong cation exchange HPLC peptide fractionation, and immobilized metal affinity chromatography prior to mass spectrometry. In total, 217 unique phosphopeptides sites were identified representing 85 distinct phosphoproteins at 2.3% global FDR. From these peptides, 129 distinct phosphorylation sites were identified of which 57 were previously known, but only 11 of which had been previously identified in whole saliva. Cellular localization analysis revealed salivary phosphoproteins had a distribution similar to all known salivary proteins, but with less relative representation in "extracellular" and "plasma membrane" categories compared to salivary glycoproteins. Sequence alignment showed that phosphorylation occurred at acidic-directed kinase, proline-directed, and basophilic motifs. This differs from plasma phosphoproteins, which predominantly occur at Golgi casein kinase recognized sequences. Collectively, these results suggest diverse functions for salivary phosphoproteins and multiple kinases involved in their processing and secretion. In all, this study should lay groundwork for future elucidation of the functions of salivary protein phosphorylation.

Original languageEnglish (US)
Pages (from-to)1728-1736
Number of pages9
JournalJournal of Proteome Research
Volume10
Issue number4
DOIs
StatePublished - Apr 1 2011

Fingerprint

Phosphorylation
Phosphoproteins
Saliva
Salivary Proteins and Peptides
Phosphotransferases
Casein Kinases
Affinity chromatography
Phosphopeptides
Peptides
Sequence Alignment
Cell membranes
Fractionation
Affinity Chromatography
Proline
Mass spectrometry
Cations
Mass Spectrometry
Ion exchange
Glycoproteins
Metals

Keywords

  • dynamic range compression
  • mass spectrometry
  • phosphoproteomics
  • whole saliva

Cite this

Large-scale phosphoproteomics analysis of whole saliva reveals a distinct phosphorylation pattern. / Stone, Matthew D.; Chen, Xiaobing; McGowan, Thomas; Bandhakavi, Sricharan; Cheng, Bin; Rhodus, Nelson L; Griffin, Timothy J.

In: Journal of Proteome Research, Vol. 10, No. 4, 01.04.2011, p. 1728-1736.

Research output: Contribution to journalArticle

Stone, Matthew D. ; Chen, Xiaobing ; McGowan, Thomas ; Bandhakavi, Sricharan ; Cheng, Bin ; Rhodus, Nelson L ; Griffin, Timothy J. / Large-scale phosphoproteomics analysis of whole saliva reveals a distinct phosphorylation pattern. In: Journal of Proteome Research. 2011 ; Vol. 10, No. 4. pp. 1728-1736.
@article{12a72a917cdf40ccad4c3b16f97737dd,
title = "Large-scale phosphoproteomics analysis of whole saliva reveals a distinct phosphorylation pattern",
abstract = "In-depth knowledge of bodily fluid phosphoproteomes, such as whole saliva, is limited. To better understand the whole saliva phosphoproteome, we generated a large-scale catalog of phosphorylated proteins. To circumvent the wide dynamic range of phosphoprotein abundance in whole saliva, we combined dynamic range compression using hexapeptide beads, strong cation exchange HPLC peptide fractionation, and immobilized metal affinity chromatography prior to mass spectrometry. In total, 217 unique phosphopeptides sites were identified representing 85 distinct phosphoproteins at 2.3{\%} global FDR. From these peptides, 129 distinct phosphorylation sites were identified of which 57 were previously known, but only 11 of which had been previously identified in whole saliva. Cellular localization analysis revealed salivary phosphoproteins had a distribution similar to all known salivary proteins, but with less relative representation in {"}extracellular{"} and {"}plasma membrane{"} categories compared to salivary glycoproteins. Sequence alignment showed that phosphorylation occurred at acidic-directed kinase, proline-directed, and basophilic motifs. This differs from plasma phosphoproteins, which predominantly occur at Golgi casein kinase recognized sequences. Collectively, these results suggest diverse functions for salivary phosphoproteins and multiple kinases involved in their processing and secretion. In all, this study should lay groundwork for future elucidation of the functions of salivary protein phosphorylation.",
keywords = "dynamic range compression, mass spectrometry, phosphoproteomics, whole saliva",
author = "Stone, {Matthew D.} and Xiaobing Chen and Thomas McGowan and Sricharan Bandhakavi and Bin Cheng and Rhodus, {Nelson L} and Griffin, {Timothy J}",
year = "2011",
month = "4",
day = "1",
doi = "10.1021/pr1010247",
language = "English (US)",
volume = "10",
pages = "1728--1736",
journal = "Journal of Proteome Research",
issn = "1535-3893",
publisher = "American Chemical Society",
number = "4",

}

TY - JOUR

T1 - Large-scale phosphoproteomics analysis of whole saliva reveals a distinct phosphorylation pattern

AU - Stone, Matthew D.

AU - Chen, Xiaobing

AU - McGowan, Thomas

AU - Bandhakavi, Sricharan

AU - Cheng, Bin

AU - Rhodus, Nelson L

AU - Griffin, Timothy J

PY - 2011/4/1

Y1 - 2011/4/1

N2 - In-depth knowledge of bodily fluid phosphoproteomes, such as whole saliva, is limited. To better understand the whole saliva phosphoproteome, we generated a large-scale catalog of phosphorylated proteins. To circumvent the wide dynamic range of phosphoprotein abundance in whole saliva, we combined dynamic range compression using hexapeptide beads, strong cation exchange HPLC peptide fractionation, and immobilized metal affinity chromatography prior to mass spectrometry. In total, 217 unique phosphopeptides sites were identified representing 85 distinct phosphoproteins at 2.3% global FDR. From these peptides, 129 distinct phosphorylation sites were identified of which 57 were previously known, but only 11 of which had been previously identified in whole saliva. Cellular localization analysis revealed salivary phosphoproteins had a distribution similar to all known salivary proteins, but with less relative representation in "extracellular" and "plasma membrane" categories compared to salivary glycoproteins. Sequence alignment showed that phosphorylation occurred at acidic-directed kinase, proline-directed, and basophilic motifs. This differs from plasma phosphoproteins, which predominantly occur at Golgi casein kinase recognized sequences. Collectively, these results suggest diverse functions for salivary phosphoproteins and multiple kinases involved in their processing and secretion. In all, this study should lay groundwork for future elucidation of the functions of salivary protein phosphorylation.

AB - In-depth knowledge of bodily fluid phosphoproteomes, such as whole saliva, is limited. To better understand the whole saliva phosphoproteome, we generated a large-scale catalog of phosphorylated proteins. To circumvent the wide dynamic range of phosphoprotein abundance in whole saliva, we combined dynamic range compression using hexapeptide beads, strong cation exchange HPLC peptide fractionation, and immobilized metal affinity chromatography prior to mass spectrometry. In total, 217 unique phosphopeptides sites were identified representing 85 distinct phosphoproteins at 2.3% global FDR. From these peptides, 129 distinct phosphorylation sites were identified of which 57 were previously known, but only 11 of which had been previously identified in whole saliva. Cellular localization analysis revealed salivary phosphoproteins had a distribution similar to all known salivary proteins, but with less relative representation in "extracellular" and "plasma membrane" categories compared to salivary glycoproteins. Sequence alignment showed that phosphorylation occurred at acidic-directed kinase, proline-directed, and basophilic motifs. This differs from plasma phosphoproteins, which predominantly occur at Golgi casein kinase recognized sequences. Collectively, these results suggest diverse functions for salivary phosphoproteins and multiple kinases involved in their processing and secretion. In all, this study should lay groundwork for future elucidation of the functions of salivary protein phosphorylation.

KW - dynamic range compression

KW - mass spectrometry

KW - phosphoproteomics

KW - whole saliva

UR - http://www.scopus.com/inward/record.url?scp=79953716847&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=79953716847&partnerID=8YFLogxK

U2 - 10.1021/pr1010247

DO - 10.1021/pr1010247

M3 - Article

VL - 10

SP - 1728

EP - 1736

JO - Journal of Proteome Research

JF - Journal of Proteome Research

SN - 1535-3893

IS - 4

ER -