Clean STD-NMR spectrum for improved detection of ligand-protein interactions at low concentration of protein

Youlin Xia, Qi Zhu, Kyu Yeon Jun, Jingchun Wang, Xiaolian Gao

Research output: Contribution to journalArticle

14 Scopus citations

Abstract

Saturation transfer difference (STD)-NMR has been widely used to screen ligand compound libraries for their binding activities to proteins and to determine the binding epitopes of the ligands. We report herein, a Clean STD-NMR method developed to overcome false positives (artifacts) observed in the STD-NMR spectrum due to the power spillover of RF irradiation. The method achieved higher degree of resonance saturation through digital editing of two STD-NMR spectra to generate a concatenated difference spectrum and three times of sensitivity enhancement for a loose binding complex involving DNA oligonucleotide and an RNA-binding protein, CUGBP-1ab (25.2 kDa). The interesting binding characteristics of the complex dCTGTCT-CUGBP1ab were obtained. The method was applied to a mixture of small ligand and bovine serum albumin protein (BSA, 66.3 kDa), and detected the intermolecular contacts at a BSA concentration as low as 0.1 μM, a working concentration useful for the detection of proteins of low solubility at biologically relevant conditions.

Original languageEnglish (US)
Pages (from-to)918-924
Number of pages7
JournalMagnetic Resonance in Chemistry
Volume48
Issue number12
DOIs
StatePublished - Dec 2010

Keywords

  • Clean STD-NMR
  • DNA-protein complex
  • STD-NMR
  • low concentration protein
  • protein binding

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