title: Ancylostoma caninum MTP-1, an astacin-like metalloprotease secreted by infective hookworm larvae, is involved in tissue migration.
abstract: Infective larvae (L3) of nematodes secrete macromolecules that are critical to infection and establishment of the parasite in the host. The dog hookworm Ancylostoma caninum secretes an astacin-like metalloprotease, Ac-MTP-1, upon activation in vitro with host serum. Recombinant Ac-MTP-1 was expressed in the baculovirus/insect cell system as a secreted protein and was purified from culture medium by two separate methods, cation-exchange fast-performance liquid chromatography and gelatin-affinity chromatography. Recombinant MTP-1 was catalytically active and digested a range of native and denatured connective tissue substrates, including gelatin, collagen, laminin, and fibronectin. A dog was immunized with recombinant Ac-MTP-1 formulated with AS03 adjuvant, and the antiserum was used to immunolocalize the anatomic sites of expression within A. caninum L3 to secretory granules in the glandular esophagus and the channels that connect the esophagus to the L3 surface and to the cuticle. Antiserum inhibited the ability of recombinant MTP-1 to digest collagen by 85% and inhibited larval migration through tissue in vitro by 70 to 75%, in contrast to just 5 to 10% inhibition obtained with preimmunization serum. The metalloprotease inhibitors EDTA and 1,10-phenanthroline also reduced the penetration of L3 through skin in vitro by 43 to 61%. The data strongly suggest that Ac-MTP-1 is critical for the invasion process of hookworm larvae, and moreover, that antibodies against the enzyme can neutralize its function and inhibit migration.
authors.affiliation: Department of Microbiology and Tropical Medicine, George Washington University Medical Center, Washington, DC, USA.
authors.forename: Angela L
authors.initials: AL
authors.lastname: Williamson
authors.forename: Sara
authors.initials: S
authors.lastname: Lustigman
authors.forename: Yelena
authors.initials: Y
authors.lastname: Oksov
authors.forename: Vehid
authors.initials: V
authors.lastname: Deumic
authors.forename: Jordan
authors.initials: J
authors.lastname: Plieskatt
authors.forename: Susana
authors.initials: S
authors.lastname: Mendez
authors.forename: Bin
authors.initials: B
authors.lastname: Zhan
authors.forename: Maria Elena
authors.initials: ME
authors.lastname: Bottazzi
authors.forename: Peter J
authors.initials: PJ
authors.lastname: Hotez
authors.forename: Alex
authors.initials: A
authors.lastname: Loukas
mesh_terms: D000722:Ancylostoma / Q000201:enzymology / Q000254:growth & development / Q000472:pathogenicity*; D000724:Ancylostomiasis / Q000469:parasitology; D000818:Animals; D003238:Connective Tissue / Q000378:metabolism / Q000469:parasitology*; D004285:Dogs; D006790:Host-Parasite Interactions; D007814:Larva / Q000201:enzymology; D008297:Male; D008666:Metalloendopeptidases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012867:Skin / Q000378:metabolism / Q000469:parasitology*
chemical_list: D011994:Recombinant Proteins; C456261:MTP-1 protein, Ancylostoma caninum; D008666:Metalloendopeptidases