Developmentally regulated secretion of cathepsin L-like cysteine proteases by Haemonchus contortus.
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.
Rhoads
M L
ML
Livestock and Poultry Sciences Institute, United States Department of Agriculture, Beltsville, Maryland 20705, USA.
Fetterer
R H
RH
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
D000871:Anthelmintics; D005456:Fluorescent Dyes; D011506:Proteins; D002403:Cathepsins; D010450:Endopeptidases; D003546:Cysteine Endopeptidases; D056668:Cathepsin L