High resolution epitope mapping of an anti-DNA autoantibody using model DNA ligands.
Although Ab-DNA complexes are involved in the pathogenesis of SLE, the mechanism by which anti-DNA recognize their DNA targets remains elusive. We have designed and synthesized small test DNA ligands that model ssDNA and dsDNA determinants commonly recognized by anti-DNA autoantibodies. These ligands were used in DNA footprinting assays to conduct a high resolution epitope-mapping study of anti-ssDNA BV04-01, which is typical of anti-ssDNA expressed in (NZB x NZW)F1 mice. We find that BV04-01 recognizes the ssDNA, but not dsDNA, epitopes presented on the test ligands, and makes specific base and backbone contacts with the DNA. Moreover, BV04-01 binding induces conformational changes in both the single-stranded and duplex region of the ligands. The epitope-mapping data is used in conjunction with computational molecular modeling to construct a three-dimensional model of a BV04-01-ligand complex. The salient aspects of this model are consistent with many features observed in the crystal structure of BV04-01 liganded to d(pT)3. This work extends previous anti-DNA binding studies and attempts to provide a unified approach that draws upon the merits of many precedented experimental methods to assemble three-dimensional structures of anti-DNA-DNA complexes. Analysis of these structures may ultimately help guide the development of molecules designed to inhibit anti-DNA binding in SLE.
Swanson
P C
PC
Department of Chemistry, University of Michigan, Ann Arbor 48109-1055.
Cooper
B C
BC
Glick
G D
GD
D000818:Animals; D000974:Antibodies, Antinuclear / Q000737:chemistry / Q000235:genetics*; D001483:Base Sequence; D001665:Binding Sites; D004247:DNA / Q000737:chemistry / Q000235:genetics* / Q000276:immunology*; D016172:DNA Fingerprinting; D004563:Electrochemistry; D000939:Epitopes / Q000235:genetics; D006801:Humans; D008024:Ligands; D008180:Lupus Erythematosus, Systemic / Q000235:genetics / Q000276:immunology; D051379:Mice; D008954:Models, Biological; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D011487:Protein Conformation
D000974:Antibodies, Antinuclear; D000939:Epitopes; D008024:Ligands; D004247:DNA