Reactivity of the antibodies to DNA modified by the carcinogen N-acetoxy-N-acetyl-2-aminofluorene

Abstract

Rabbits were immunized with native DNA modified by the carcinogen N-acetoxy-N-acetyl-2-aminofluorene. The interactions between the purified antibodies to nDNA-AAF (or the Fab fragments) and several ligands were studied. By radioimmunoassay, nDNA-AAF, dDNA-AAF, and GMP-AAF were found to bind to the antibodies with about the same affinity. GMP-AF interacts slightly less, and GMP and N-OH-AAF do not interact. The values of the association constants deduced from fluorescence measurements for the binding of the Fab fragments to nDNA-AAF, dDNA-AAF, and GMP-AAF, in 50 mM NaCl, pH 7.5, are of the same order of magnitude. The values of the association constants with nDNA-AAF and dDNA-AAF depend on salt concentration. From this variation, it is deduced that 1-1.5 phosphate groups interact by charge-charge interactions with the Fab fragments. The absorption and circular dichroism spectra of GMP-AAF, nDNA-AAF, and dDNA-AAF bound to the Fab fragments show that the Fab fragments induce similar perturbation to the 3 ligands. The immunodeterminant group appears to be the dGMP-AAF residue.