title: Electron spin resonance spin-label studies of intercalation of ethidium bromide and aromatic amine carcinogens in DNA.
abstract: A spin-labeled technique has been developed for the study of carcinogen-DNA intercalation mechanisms utilizing a modified ethidium bromide as a model ligand and specifically synthesized spin labels of carcinogenic aromatic amines such as 2-aminofluorene, 2-aminoanthracene, and 6-aminochrysene. The technique takes advantage of the characteristic electron spin resonance anisotropy of the corresponding spin labels complexed with DNA. Analysis of the orientation-dependent information thus obtained has clearly shown that the lignad moieties in all 4 respective complexes are accommodated in adjacent base-pair layers of the DNA with the nitroxide reporter oriented in 1 preferential direction. The geometry of the nitroxide rings with respect to their respective constraining ligands in a particular complex appeared to be largely dependent upon the size of the ligand moiety and the position of the reporter attachment on the ligand molecule. A coplanar configuration is most favored for the spin labels in which the ligand moiety per se is a 3-fused ring with the nitroxide substituted at a position on or close to the longitudinal end of the ligand molecule. Temperature studies in which the heat-induced release of the bound label is monitored by electron spin resonance gives an accurate measure of the complex dissociation characteristics that appear to parallel the helical coil cooperative melting transition monitored by absorbance measurements. The extent of dissociation of the bound spin labels parallels almost exactly the melting, i.e., unzipping of the DNA double helix, suggesting that the spin labels are immobilized by intercalation at the site on the DNA directly involved in the maintenance of the double helical structure. Analysis of the electron spin resonance spectra of the complexes in terms of the extent of immobilization indicates that, for aromatic amine spin labels, two different binding species exist, one strongly immobilized presumably at a guanine-cytosine base-pair region and the other partially immobilized in a region other than guanine-cytosine base pair. Competitive binding of the labels with respect to their parent ligand and binding isotherm results show that nitroxide substitution does not affect the type of binding but does affect the extent of binding, i.e.. binding constant with the labeled compound having a lower affinity.
mesh_terms: D000588:Amines; D000873:Anthracenes / Q000032:analysis; D002273:Carcinogens / Q000032:analysis*; D004247:DNA / Q000032:analysis* / Q000378:metabolism; D004996:Ethidium; D005449:Fluorenes / Q000032:analysis; D008024:Ligands; D008956:Models, Chemical; D010616:Phenanthrenes / Q000032:analysis; D013113:Spin Labels
chemical_list: D000588:Amines; D000873:Anthracenes; D002273:Carcinogens; D005449:Fluorenes; D008024:Ligands; D010616:Phenanthrenes; D013113:Spin Labels; C012177:2-aminofluorene; C006593:2-anthramine; D004247:DNA; D004996:Ethidium