Site-specific DNA-affinity chromatography of the lac repressor

Nucleic Acids Res. 1980 Aug 25;8(16):3721-8. doi: 10.1093/nar/8.16.3721.

Abstract

To test the feasibility of site-specific DNA-affinity chromatography, E. coli lac repressor was bound to an operator-containing DNA column, and in parallel to a non-operator DNA column. Salt gradient elution shows: 1) elution from non-operator DNA was near 250mM KCl or NaCl; interpretation of this result suggests the usefulness of the procedure for studying salt-dependence of DNA-protein affinities; 2) elution from operator-containing DNA was delayed (average elution = 1000mM salt), demonstrating a feasibility of site-specific DNA-affinity chromatography, if one provides a sufficiently favorable ratio of specific to non-specific DNA binding sites; 3) repressor eluted from operator-containing DNA over a very broad salt range, which may represent chromatography-generated repressor heterogeneity.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Chromatography, Affinity
  • DNA, Bacterial
  • Escherichia coli / analysis*
  • Kinetics
  • Plasmids
  • Repressor Proteins / isolation & purification*
  • Transcription Factors / isolation & purification*

Substances

  • DNA, Bacterial
  • Repressor Proteins
  • Transcription Factors