Specificity of autoimmune monoclonal Fab fragments binding to single-stranded deoxyribonucleic acid.
Fab fragments from hybridoma HEd 10 [Lee, J. S., Lewis, J.R., Morgan, A.R., Mosmann, T.R., & Singh, B. (1981) Nucleic Acids Res. 9, 1707-1721] were prepared in large amounts by papain digestion of the immunoglobulin G (IgG) fraction from ascites fluid. Binding data were generated by a fluorescence quenching technique, and binding constants [K(0)] were estimated from Scatchard plots. The Fab fragments bound tightly to poly(dT) [K(0) = 12.7 X 10(6) M-1], and analysis of binding constants for the series p(dT)2 to p(dT)17 showed that the recognition sequence consisted of four consecutive residues. The effect of ionic strength on the interaction suggested that only two phosphates were involved. Binding constants for poly(dU), poly[d(brU)], poly[d(brC)], and poly(rU) were 1.0 X 10(6) M-1, 18.8 X 10(6) M-1, 0.5 X 10(6) M-1, and less than 0.5 X 10(6) M-1, respectively, implicating the involvement of the 3, 4, and 5 positions of the pyrimidine ring as well as the deoxyribose sugar in the recognition process. At high ionic strength (0.5 M) K(0) for whole IgG binding to poly(dT) was 75 X 10(6) M-1 compared to a value of 1.1 X 10(6) M-1 for the Fab fragment. These results may have implications for the tissue damage caused by DNA-containing immune complexes in systemic lupus erythematosus.
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D001323:Autoantibodies / Q000276:immunology; D001483:Base Sequence; D001666:Binding Sites, Antibody; D002460:Cell Line; D004277:DNA, Single-Stranded / Q000276:immunology*; D000939:Epitopes; D006825:Hybridomas; D007140:Immunoglobulin Fab Fragments / Q000276:immunology*; D051379:Mice; D008815:Mice, Inbred Strains
D000911:Antibodies, Monoclonal; D001323:Autoantibodies; D004277:DNA, Single-Stranded; D000939:Epitopes; D007140:Immunoglobulin Fab Fragments