title: Distamycin A affects the stability of NF-kappaB p50-DNA complexes in a sequence-dependent manner.
abstract: The effect of two different DNA minor groove binding molecules, Hoechst 33258 and distamycin A, on the binding kinetics of NF-kappaB p50 to three different specific DNA sequences was studied at various salt concentrations. Distamycin A was shown to significantly increase the dissociation rate constant of p50 from the sequences PRDII (5'-GGGAAATTCC-3') and Ig-kappa B (5'-GGGACTTTCC-3') but had a negligible effect on the dissociation from the palindromic target-kappaB binding site (5'-GGGAATTCCC-3'). By comparison, the effect of Hoechst 33258 on binding of p50 to each sequence was found to be minimal. The dissociation rates for the protein--DNA complexes increased at higher potassium chloride concentrations for the PRDII and Ig-kappaB binding motifs and this effect was magnified by distamycin A. In contrast, p50 bound to the palindromic target-kappaB site with a much higher intrinsic affinity and exhibited a significantly reduced salt dependence of binding over the ionic strength range studied, retaining a K(D) of less than 10 pM at 150 mM KCl. Our results demonstrate that the DNA binding kinetics of p50 and their salt dependence is strongly sequence-dependent and, in addition, that the binding of p50 to DNA can be influenced by the addition of minor groove-binding drugs in a sequence-dependent manner.
authors.affiliation: Department of Biochemistry, University of Cambridge, Tennis Court Road, Cambridge CB2 1GA, UK.
authors.forename: Robert E
authors.initials: RE
authors.lastname: Speight
authors.forename: Darren J
authors.initials: DJ
authors.lastname: Hart
authors.forename: Jonathan M
authors.initials: JM
authors.lastname: Blackburn
mesh_terms: D001483:Base Sequence; D001562:Benzimidazoles; D001665:Binding Sites / Q000235:genetics; D004247:DNA / Q000235:genetics / Q000378:metabolism*; D004214:Distamycins / Q000378:metabolism*; D007700:Kinetics; D016328:NF-kappa B / Q000378:metabolism*; D011485:Protein Binding
chemical_list: D001562:Benzimidazoles; D004214:Distamycins; D016328:NF-kappa B; C447680:hoechst 32258; C030000:stallimycin; D004247:DNA