Strong anion exchange for studying protein-DNA interactions by H/D exchange mass spectrometry

J Am Soc Mass Spectrom. 2008 Jun;19(6):887-90. doi: 10.1016/j.jasms.2008.03.003. Epub 2008 Mar 22.

Abstract

The use of mass spectrometry to study protein-ligand interactions is expanding into more complex systems including protein-DNA interactions. The excess amount of a model DNA or, more typically, an oligodeoxynucleotide (ODN), needed to study such interactions in an amide hydrogen-deuterium (H/D) exchange experiment, for example, causes serious signal suppression in the protein analysis. We describe here a modification of the traditional H/D exchange protocol whereby we utilize a strong anion exchange column to rapidly remove the ODN from solution before MS analysis. We showed the successful incorporation of such a column in a study of two protein-ODN systems: (1) the DNA-binding domain of human telomeric repeat binding factor 2 with a telomeric oligodeoxynucleotide and (2) thrombin with the thrombin-binding aptamer. The approach gave no appreciable difference in back-exchange compared to a method in which no strong anion exchange (SAX) is used.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Anions
  • Binding Sites
  • Chromatography, Ion Exchange / methods*
  • DNA / chemistry*
  • DNA-Binding Proteins / chemistry*
  • Deuterium Exchange Measurement / methods*
  • Protein Binding
  • Protein Interaction Mapping / methods*
  • Spectrometry, Mass, Electrospray Ionization / methods*

Substances

  • Anions
  • DNA-Binding Proteins
  • DNA