EhCP112 is an Entamoeba histolytica secreted cysteine protease that may be involved in the parasite-virulence.
EhCP112 is an Entamoeba histolytica protease that together with the EhADH112 protein forms the EhCPADH complex involved in trophozoite virulence. Here, we produced the recombinant EhCP112 and studied its relationships with extracellular matrix components and with target cells. A DNA fragment containing the pro-peptide and the mature enzyme was expressed in bacteria as an active enzyme (rEhCP112), whereas the full gene containing the signal peptide, the pro-peptide and the mature enzyme expressed a non-active protein. The fragment only with the mature enzyme was not expressed. rEhCP112 purified by affinity columns digested azocasein and had a strong autoproteolytic activity. Four hours after purification the protein appeared degraded. Anti-tag antibodies, monoclonal antibodies against the EhCP112 and sera from human patients with amoebiasis recognized rEhCP112. rEhCP112 digested gelatin, collagen type I, fibronectin and haemoglobin; it destroyed MDCK cell monolayers and bound to red blood cells. The native EhCP112 was poorly expressed in a virulence-deficient mutant, and in the wild-type clone it was located in secreted vesicles, forming the EhCPADH complex. Altogether these results show that EhCP112 is a molecule able to disrupt cell monolayers and digest proteins of the extracellular matrix and haemoglobin, and it is secreted by the trophozoites.
Ocádiz
Ramón
R
Escuela Nacional de Medicina y Homeopatía, IPN, Guillermo Massieu Helguera #239, México, DF, 07320.
Orozco
Esther
E
Carrillo
Eduardo
E
Quintas
Laura Itzel
LI
Ortega-López
Jaime
J
García-Pérez
Rosa María
RM
Sánchez
Tomas
T
Castillo-Juárez
Beatriz A
BA
García-Rivera
Guillermina
G
Rodríguez
Mario A
MA
D000818:Animals; D000913:Antibodies, Protozoan / Q000276:immunology; D001426:Bacterial Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D002364:Caseins / Q000378:metabolism; D003001:Cloning, Molecular; D024042:Collagen Type I / Q000378:metabolism; D006224:Cricetinae; D003546:Cysteine Endopeptidases / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D004748:Entamoeba histolytica / Q000201:enzymology* / Q000235:genetics / Q000472:pathogenicity; D004749:Entamoebiasis / Q000276:immunology / Q000469:parasitology; D004912:Erythrocytes / Q000469:parasitology; D005109:Extracellular Matrix / Q000378:metabolism; D005353:Fibronectins / Q000378:metabolism; D005780:Gelatin / Q000378:metabolism; D006454:Hemoglobins / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D008297:Male; D051379:Mice; D009154:Mutation; D011817:Rabbits; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism
D000913:Antibodies, Protozoan; D001426:Bacterial Proteins; D002364:Caseins; D024042:Collagen Type I; D005353:Fibronectins; D006454:Hemoglobins; D011994:Recombinant Proteins; C026018:azocasein; D005780:Gelatin; D003546:Cysteine Endopeptidases; C122004:EhCP112 protein, Entamoeba histolytica