| 1559 Proline-rich Proteins as Major Part of the Human Saliva Proteome | ||
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A. WALZ1, A. WATTENBERG2, G. SCHMALZ3, M. BLUEGGEL2, and S. RUHL3, 1University of Regensburg, Dental School, Germany, 2Protagen AG, Dortmund, Germany, 3University of Regensburg, Germany Proline-rich proteins (PRPs) are the major group of proteins in human saliva. Although proteome analysis has been performed on human saliva, no positive identification of proline-rich proteins by a combination of 2D-gelelectrophoresis (2-DE) and mass spectrometry (MS) is available. Objective: The aim of this study was to enlarge the two-dimensional protein map of whole human saliva by including the display of proline-rich proteins in 2D gels and their subsequent identification by mass spectrometry. Methods: Both whole saliva proteins and purified PRP-1 were subjected to isoelectric focusing followed by SDS-PAGE over a pH range between 3-10. Spots were picked, digested, and analyzed by MALDI (matrix assisted laser desorption/ionization)-MS. The data generated were analyzed using protein data base search. Results: New components identified by 2-DE and MS in saliva were actin, cyclophilin B, phosphoglycerate kinase, prolyl-4-hydroxylase, thioredoxin and transferrin. After deviating from the usual database search parameters with regard to trypsin specificity, the identity of PRP-1 could be confirmed by MALDI TOF (time of flight) and MALDI-TOF/TOF. Based on this, an identification of PRPs in saliva was achieved. Interestingly, the proline-rich proteins could only be displayed in 2D-gels when saliva was filtrated with a membrane filter of 0,2 µm pore size. Among the 130 spots identified, 30 spots were acidic proline-rich phosphoproteins, one was proline-rich protein BstNI, subfamiliy 1, and one was proline-rich protein BstNI, subfamily 3. Concerning the acidic proline-rich phosphoproteins an additional peptide peak with a mass of 2136 kDa was detected that was not present in purified PRP-1. The amino acid sequence of this peptide as revealed by peptide-fragmentation-fingerprint (PFF) identified PRP Db (gene PRH1) in saliva. Conclusion: Proline-rich proteins can be displayed by 2D-PAGE and represent, besides salivary amylase, the major part of human salivary proteome. Supported by grant SFB 585/B5 from the DFG | ||
| Seq #192 - Salivary Proteomics 10:45 AM-12:45 PM, Friday, 11 March 2005 Baltimore Convention Center 322-323 | ||
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