Large-scale phosphoproteome analysis of human liver tissue by enrichment and fractionation of phosphopeptides with strong anion exchange chromatography.

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Han G, Ye M, Zhou H, Jiang X, Feng S, Jiang X, Tian R, Wan D, Zou H, Gu J

Large-scale phosphoproteome analysis of human liver tissue by enrichment and fractionation of phosphopeptides with strong anion exchange chromatography.

Proteomics. 2008 Apr;8(7):1346-61. doi: 10.1002/pmic.200700884.

PubMed ID
18318008 [ View in PubMed
]
Abstract

The mixture of phosphopeptides enriched from proteome samples are very complex. To reduce the complexity it is necessary to fractionate the phosphopeptides. However, conventional enrichment methods typically only enrich phosphopeptides but not fractionate phosphopeptides. In this study, the application of strong anion exchange (SAX) chromatography for enrichment and fractionation of phosphopeptides was presented. It was found that phosphopeptides were highly enriched by SAX and majority of unmodified peptides did not bind onto SAX. Compared with Fe(3+) immobilized metal affinity chromatography (Fe(3+)-IMAC), almost double phosphopeptides were identified from the same sample when only one fraction was generated by SAX. SAX and Fe(3+)-IMAC showed the complementarity in enrichment and identification of phosphopeptides. It was also demonstrated that SAX have the ability to fractionate phosphopeptides under gradient elution based on their different interaction with SAX adsorbent. SAX was further applied to enrich and fractionate phosphopeptides in tryptic digest of proteins extracted from human liver tissue adjacent to tumorous region for phosphoproteome profiling. This resulted in the highly confident identification of 274 phosphorylation sites from 305 unique phosphopeptides corresponding to 168 proteins at false discovery rate (FDR) of 0.96%.

DrugBank Data that Cites this Article

Polypeptides
NameUniProt ID
Ferritin heavy chainP02794Details
AlbuminP02768Details
Ryanodine receptor 1P21817Details
5'-AMP-activated protein kinase subunit beta-2O43741Details
Thiopurine S-methyltransferaseP51580Details
CalnexinP27824Details
Heat shock protein HSP 90-alphaP07900Details
FibronectinP02751Details
Heat shock protein HSP 90-betaP08238Details
Histone deacetylase 1Q13547Details
Prostaglandin E synthase 3Q15185Details
cAMP-dependent protein kinase type II-alpha regulatory subunitP13861Details
cAMP-dependent protein kinase type I-alpha regulatory subunitP10644Details
Cation-independent mannose-6-phosphate receptorP11717Details
Proteasome subunit alpha type-3P25788Details
Calcium/calmodulin-dependent protein kinase type II subunit deltaQ13557Details
Myosin-11P35749Details
cAMP-dependent protein kinase type II-beta regulatory subunitP31323Details
14-3-3 protein epsilonP62258Details
Membrane-associated progesterone receptor component 1O00264Details
Nucleolar and coiled-body phosphoprotein 1Q14978Details
Hepatoma-derived growth factorP51858Details
Protein S100-A9P06702Details
Alpha-2-HS-glycoproteinP02765Details
Heterogeneous nuclear ribonucleoprotein D0Q14103Details
NucleophosminP06748Details
Myosin-9P35579Details
Myosin light chain kinase, smooth muscleQ15746Details
Serine/threonine-protein kinase OSR1O95747Details