An immunodominant epitope on DNA topoisomerase I is conformational in nature: heterogeneity in its recognition by systemic sclerosis sera.
OBJECTIVE
To characterize an immunodominant epitope recognized by anti-DNA topoisomerase I (topo I) antibody, a major autoantibody in sera of patients with systemic sclerosis (SSc).


METHODS
Topo I fragments were generated as fusion proteins using a bacterial expression system as well as polypeptides translated in vitro using a eukaryotic expression system. Reactivities to the 2 preparations of recombinant topo I polypeptides in anti-topo I-positive sera from SSc patients of varied ethnic backgrounds were examined by immunoblotting, immunoprecipitation, and/or enzyme-linked immunosorbent assay.


RESULTS
The fragment encoding amino acids 489-573 of topo I was recognized by 98 of 100 anti-topo I-positive SSc sera. Both carboxyl- and amino-terminal deletion studies as well as competitive inhibition assays using topo I synthetic peptides showed that a region of > or =52 amino acids (512-563) was necessary for recognition by anti-topo I antibodies. The minimum epitope region and conformation required for this reactivity were variable among sera from Caucasian, African American, Japanese, and Choctaw SSc patients.


CONCLUSIONS
An immunodominant epitope recognized by anti-topo I autoantibody is located in the region of amino acids 489-573 of the topo I protein and is largely conformational in nature. The recognition pattern of this region by anti-topo I-positive sera is heterogeneous and is influenced by ethnic background.
Kuwana
M
M
University of Pittsburgh School of Medicine, Pennsylvania 15213, USA.
Kaburaki
J
J
Medsger
T A
TA
Wright
T M
TM
D000595:Amino Acid Sequence; D001323:Autoantibodies / Q000276:immunology*; D001324:Autoantigens / Q000032:analysis*; D004264:DNA Topoisomerases, Type I / Q000032:analysis* / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay; D005006:Ethnicity; D000949:Histocompatibility Antigens Class II / Q000235:genetics; D006801:Humans; D015151:Immunoblotting; D016056:Immunodominant Epitopes / Q000032:analysis*; D008969:Molecular Sequence Data; D011233:Precipitin Tests; D011993:Recombinant Fusion Proteins / Q000032:analysis; D012595:Scleroderma, Systemic / Q000235:genetics / Q000276:immunology*
D001323:Autoantibodies; D001324:Autoantigens; D000949:Histocompatibility Antigens Class II; D016056:Immunodominant Epitopes; D011993:Recombinant Fusion Proteins; D004264:DNA Topoisomerases, Type I