title: Developmentally regulated secretion of cathepsin L-like cysteine proteases by Haemonchus contortus.
abstract: Cysteine protease activity was present in media collected after 24 hr in vitro culture of adult Haemonchus contortus. The released cysteine protease hydrolyzed the fluorogenic 7-amino-4-trifluoromethyl coumarin (AFC)-substituted synthetic peptides Z-phe-arg-AFC and Z-ala-arg-arg-AFC, but not Z-arg-arg-AFC or Z-arg-AFC, characterizing this activity as cathepsin L-like. Within the parasite, cysteine protease activity was highest in extracts of intestinal tissue. Secreted cysteine protease inhibited the clotting of sheep blood and hydrolyzed hemoglobin, fibrinogen, collagen, and IgG; the IgG hydrolysis site was within the hinge region. Four proteases with M(r) values of 30, 34, 37, and 41 kDa were identified with biotinylated-phenylalanine-arginine-fluoromethyl ketone, a specific probe that binds to active cysteine proteases. Adult parasites cultivated in the presence of 0.1 mM levamisole released 50% less protease activity compared to control cultures; in the presence of rafoxanide (0.1 mM), protease was not detected. Cathepsin L-like cysteine protease activity was released also by L4, but not the L3 larval stage. The active and developmentally regulated release of cysteine proteases by H. contortus may have a critical function in worm nutrition, immune evasion, or both.
authors.affiliation: Livestock and Poultry Sciences Institute, United States Department of Agriculture, Beltsville, Maryland 20705, USA.
authors.forename: M L
authors.initials: ML
authors.lastname: Rhoads
authors.forename: R H
authors.initials: RH
authors.lastname: Fetterer
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000871:Anthelmintics / Q000494:pharmacology; D001777:Blood Coagulation; D056668:Cathepsin L; D002403:Cathepsins / Q000378:metabolism*; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases; D005456:Fluorescent Dyes / Q000737:chemistry / Q000378:metabolism; D006189:Haemonchus / Q000187:drug effects / Q000201:enzymology*; D006868:Hydrolysis; D007814:Larva / Q000201:enzymology; D008969:Molecular Sequence Data; D011506:Proteins / Q000378:metabolism; D012756:Sheep; D013379:Substrate Specificity
chemical_list: D000871:Anthelmintics; D005456:Fluorescent Dyes; D011506:Proteins; D002403:Cathepsins; D010450:Endopeptidases; D003546:Cysteine Endopeptidases; D056668:Cathepsin L