Isolation, characterization and expression of a GRA2 homologue from Neospora caninum

A cDNA library derived from mRNA of tachyzoites of Neospora caninum (NC-Liv erpool strain) was screened with antisera from a cow naturally infected wit h N. caninum. The DNA sequence of 1 recombinant isolated predicted a signif icant protein sequence homology of the gene product to the 28 kDa (GRA2) an tigen of Toxoplasma gondii. Studies on the N. caninum gene coding for this antigen demonstrated the presence of a single intron flanked by 2 exons; th e gene was also highly expressed in culture-derived tachyzoites. The antige n was expressed in Escherichia coli; when injected into mice it stimulated the production of antibodies which detected a 29 kDa antigen of N. caninum. Secondary structure predictions made for the N. caninum protein showed sup port for several amphipathic helices separated by loops and turns. The avai lable evidence indicates maintenance of protein secondary structure, and no t DNA or amino acid sequence, has occurred during the evolution of GRA2 pro teins in N. caninum and T. gondii.