Analysis of Antigen Binding Activity and Enzyme Inhibitory Activity of lgG Antibody to DNA Topoisomerase I from Patients with Collagen Diseases.

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The antigen of anti-Og antibody which had been found by the auther independently in Japan in a patient named Ogino with progressive systemic sclerosis has been clarified to be DNA topoisomerase I. To get a better understanding of the antigen antibody system, antibody positive lgG was purified and analysed for antigen binding activity and enzyme inhibitory activity to DNA topoisomerase I (topo I).Antinuclear antibody positive sera were first sereened by indirect immunofluorescent test. Then precipitating antibodies to DNA topoisomerase I were screened in fluorescent antinuclear antibody positive sera by double immunodiffusion test using rabbit thymus extracts as the antigen. Through these screening tests 72 sera containing anti-topo I antibody could be collected in 261 sefa from patients with PSS. By using immunoblotting technique and enzyme immunosorbent assay (ELISA), these sera were confirmed to contain IgG antibody to topo I. IgG fractions were separated by using DE52 column chromatography from 17 sera containing IgG antibodies. Antigen binding activities of the separated IgG samples were determined simultaneously by ELISA using affinity-purified topo I from rabbit thymus extract as the antigen. Enzyme inhibitory activity of the separated IgG samples were assayed by determining the minimal dosis of IgG to inhibit the relaxation of supercoiled DNA (pBR322 plasmid DNA) by calf thymus DNA topoisomerase I enzyme. The antigen binding activities of IgG samples did not correlate well to the enzyme inhibitory activities.There data indicate that the antigenic epitopes of DNA topoisomerase I are heterogeneous including the active center of the enzyme.