Site-specific DNA-affinity chromatography of the lac repressor.
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.
D002846:Chromatography, Affinity; D004269:DNA, Bacterial; D004926:Escherichia coli / Q000032:analysis*; D007700:Kinetics; D010957:Plasmids; D012097:Repressor Proteins / Q000302:isolation & purification*; D014157:Transcription Factors / Q000302:isolation & purification*
D004269:DNA, Bacterial; D012097:Repressor Proteins; D014157:Transcription Factors