Differential recognition of Toxoplasma gondii recombinant nucleoside triphosphate hydrolase isoforms by naturally infected human sera

Toxoplasma gondii possesses a highly active nucleoside triphosphate hydrola se, which has been shown to be an immunodominant antigen in mice and humans . Two isoforms (I and II) which exhibit different activities with respect t o hydrolysis of ATP exist. Past studies suggest that all strains of T. gond ii contain the less active nucleoside triphosphate hydrolase II, whilst onl y virulent strains contain the nucleoside triphosphate hydrolase I isoform. In order to further investigate the correlation between nucleoside triphos phate hydrolase isoform and biological significance, we cloned and expresse d as glutathione S-transferase fusion proteins the full-length nucleoside t riphosphate hydrolase I and II isoforms and two truncations of the nucleosi de triphosphate hydrolase I isoform in Escherichia coli. We then used ELISA s with the full-length recombinant nucleoside triphosphate hydrolases as an tigens to examine 188 naturally infected T. gondii-positive sera and 83 T. gondii-negative sera for antibody reactivity. All positive sera reacted to T. gondii whole tachyzoite lysate antigen, 31 sera reacted to both nucleosi de triphosphate hydrolase isoforms, three sera reacted specifically to nucl eoside triphosphate hydrolase I and two sera reacted to only nucleoside tri phosphate hydrolase II. Immunoblot analysis of the five sera reacting to ei ther nucleoside triphosphate hydrolase I or II revealed both quantitative a nd qualitative differences in reactivity to the two isoforms, Comparative i mmunoblot analysis using the truncations of the nucleoside triphosphate hyd rolase I isoform, and one of these positive sera identified a presumptive d ifferential epitope between the nucleoside triphosphate hydrolase I and II isoforms within an 81 amino acid region (amino acids 445-526) at the C-term inus of the nucleoside triphosphate hydrolase I isoform. This differential reactivity was further localised to the 12-residue region of greatest varia bility between the two isoforms (residues 488-499) using synthetic peptides , This is the first report where naturally infected human sera have been us ed to identify a differential epitope. Because this region is essential for substrate binding, an antibody response to this region may play some role in inhibition of this highly active enzyme. (C) 1999 Australian Society for Parasitology Inc. Published by Elsevier Science Ltd. All rights reserved.