Cloning and expression of a trypomastigote-specific 85-kilodalton surface antigen gene from Trypanosoma cruzi

An 85-kDa trypomastigote-specific surface antigen gene from Trypanosoma cruzi has been identified by screening a genomic library in λgt10 with trypomastigote and epimastigote cDNA. The 1.3-kb genomic clone (pTt34) hybridises to a single trypomastigote mRNA of 3.7 kb and to multiple bands in genomic Southern blots. Dot-blot experiments show that there are 5–10 copies of this sequence per haploid genome, and these are arranged in a non-tandem manner, pTt34 has been expressed as an anthranilate synthetase fusion protein in Escherichia coli, and inclusion bodies have been used to raise antiserum in rabbits. This antiserum immunoprecipitates a cell surface trypomastigote-specific protein of 85 kDa. The DNA and predicted amino acid sequences of pTt34 are given. Four further clones obtained from a PvuII/ HpaI partial genomic library in pUC13 have extended the sequence of the 3′ end of pTt34; each of these clones has regions of sequence divergence and each could represent a different member of the gene family.