title: Cloning, expression and characterization of immunogenic aminopeptidase N from Brucella melitensis.
abstract: A 97-kDa purified aminopeptidase N (PepN) of Brucella melitensis was previously identified to be immunogenic in humans. The B. melitensis pepN gene was cloned, expressed in Escherichia coli and purified by affinity chromatography. The recombinant PepN (rPepN) exhibited the same biochemical properties, specificity and susceptibility to inhibitors as the native PepN. rPepN was evaluated as a diagnostic antigen in an indirect enzyme-linked immunosorbent assay (ELISA) using sera from patients with acute and chronic brucellosis. The specificity of the ELISA was determined with sera from healthy donors. The ELISA had a cutoff value of 0.156 with 100% specificity and 100% sensitivity. Higher sensitivity was obtained using rPepN compared with crude extract from B. melitensis. Anti-PepN sera did not exhibit serological cross-reaction to crude extracts from Rhizobium tropici, Ochrobactrum anthropi, Yersinia enterocolitica 09 or E. coli O157H7.
authors.affiliation: Escuela Nacional de Ciencias Biológicas, IPN, México.
authors.forename: Araceli
authors.initials: A
authors.lastname: Contreras-Rodriguez
authors.forename: Mohamed Naguieb
authors.initials: MN
authors.lastname: Seleem
authors.forename: Gerhardt G
authors.initials: GG
authors.lastname: Schurig
authors.forename: Nammalwar
authors.initials: N
authors.lastname: Sriranganathan
authors.forename: Stephen M
authors.initials: SM
authors.lastname: Boyle
authors.forename: Ahide
authors.initials: A
authors.lastname: Lopez-Merino
mesh_terms: D017347:Brucella melitensis / Q000201:enzymology* / Q000235:genetics / Q000276:immunology; D002006:Brucellosis / Q000097:blood / Q000382:microbiology; D018826:CD13 Antigens / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification; D003001:Cloning, Molecular; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D006801:Humans; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification
chemical_list: D011994:Recombinant Proteins; D018826:CD13 Antigens