Combinatorial peptide ligand libraries for urine proteome analysis: Investigation of different elution systems

Giovanni Candiano, Veronica Dimuccio, Maurizio Bruschi, Laura Santucci, Rosanna Gusmano, Egisto Boschetti, Pier Giorgio Righetti, Gian Marco Ghiggeri

Research output: Contribution to journalArticle

Abstract

Proteome treatments with peptide libraries in view of reducing high-abundance proteins and increasing the concentration of rare species involve the adsorption on solid-phase material. Subsequent elution of captured proteins may not be fully effective except when sequences of eluting agents are used. The standard way utilized up to the present has been a three- to four-step, sequential elution system consisting of various agents mixed together such as urea, thiourea, CHAPS, sodium chloride, citric or acetic acid and some polar solvents such as ACN and isopropanol. Elution sequences produce distinct fractions adding to the burden of having to analyze all of them. An alternative, highly effective, single elution to reduce the workload is here reported for the first time, namely elution in boiling 10% SDS added with 3% DTE. This single step elutes almost quantitatively the adsorbed proteins, thus ensuring, for all practical purposes, a full recovery. This high efficiency is believed to be due to the fact that the SDS micelles bury the polypeptide chains within their hydrophobic core, thus shielding them from the surroundings and impeding accidental adsorption to surfaces. Suggestions for selecting the best method to eliminate the excess of SDS for further protein analysis are also evaluated. The merits and limits of this novel system are assessed and discussed.

Original languageEnglish
Pages (from-to)2405-2411
Number of pages7
JournalElectrophoresis
Volume30
Issue number14
DOIs
Publication statusPublished - Jul 2009

Keywords

  • Hexapeptide ligands
  • Human urine
  • Low-abundance proteome
  • Peptide libraries
  • SDS-elution

ASJC Scopus subject areas

  • Biochemistry
  • Clinical Biochemistry

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