Identification of proteins bound to a thioaptamer probe on a proteomics array

He Wang, Xianbin Yang, Gavin C. Bowick, Norbert K. Herzog, Bruce A. Luxon, Lee O. Lomas, David G. Gorenstein

Research output: Contribution to journalArticle

19 Scopus citations

Abstract

A rapid method to screen and identify unknown bound proteins to specific nucleic acid probes anchored on ProteinChip array surfaces from crude biological samples has been developed in this paper. It was demonstrated with screening specific binding proteins from LPS-stimulated mouse 70Z/3 pre-B cell nuclear extracts by direct coupling of thioaptamer XBY-S2 to the pre-activated ProteinChip array surfaces. With pre-fractionation of crude nuclear extracts by ion exchange method, specific "on-chip" captured proteins have been obtained that were pure enough to do "on-chip" digestion and the subsequent identification of the "on-chip" bound proteins by microsequencing of the trypsin digested peptide fragments through tandem MS. Five mouse heterogeneous nuclear ribonucleoproteins (hnRNPs) A1, A2/B1, A3, A/B, and D0 were identified. To verify those bound hnRNPs, a novel thioaptamer/antibody sandwich assay provides highly sensitive and selective identification of proteins on ProteinChip arrays.

Original languageEnglish (US)
Pages (from-to)586-593
Number of pages8
JournalBiochemical and Biophysical Research Communications
Volume347
Issue number3
DOIs
StatePublished - Sep 1 2006

Keywords

  • Aptamer array
  • DNA-binding
  • Fractionation
  • Proteomics
  • SELDI-TOF-MS
  • Sandwich assay
  • Thioaptamer
  • Transcription factor AP-1
  • hnRNPs

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

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  • Cite this

    Wang, H., Yang, X., Bowick, G. C., Herzog, N. K., Luxon, B. A., Lomas, L. O., & Gorenstein, D. G. (2006). Identification of proteins bound to a thioaptamer probe on a proteomics array. Biochemical and Biophysical Research Communications, 347(3), 586-593. https://doi.org/10.1016/j.bbrc.2006.06.132