TAENIA-SAGINATA ONCOSPHERE EXCRETORY SECRETORY PEPTIDASES/

To identify oncosphere excretory/secretory peptidases, Taenia saginata adult worms were collected from 3 patients. Eggs were hatched and act ivated in vitro and oncospheres cultured in vitro. The culture medium from the oncospheres was assayed with peptide substrates coupled to 7- amino-4-trifluoromethyl coumarin (AFC), and free AFC was detected fluo rometrically. The endopeptidase substrates Z-Phe-Arg-AFC and Z-Arg-AFC as well as the aminopeptidase substrate Arg-AFC were hydrolyzed when incubated with spent media from the oncospheres compared to control cu lture medium removed prior to incubation. Hydrolysis of Z-Phe-Arg-AFC was inhibited 78% by preincubation of the medium with the serine prote inase inhibitor Phenylmethylsulfonyl fluoride. Endopeptidase activity was partially enhanced in the presence of exogenous thiols and partial ly inhibited with the cysteine proteinase inhibitor E-64, suggesting t he presence of both serine and cysteine endopeptidases. No significant inhibition was noted with pepstatin or phenanthroline. The peptidase activities detected with Z-Phe-Arg-AFC and Arg-AFC were separated by g el-filtration fast protein liquid chromatography and eluted at volumes corresponding to molecular weights of 18 and 30 kDa, respectively. Th ese data demonstrate that T. saginata oncospheres produce excretory/se cretory peptidases, including serine and cysteine endopeptidases and a n aminopeptidase. These enzymes may play a role in invasion of the int estinal mucosa of the intermediate host.