title: Leishmania infantum: gene cloning of the GRP94 homologue, its expression as recombinant protein, and analysis of antigenicity.
abstract: The complete nucleotide sequence for the Leishmania infantum homologue to the glucose-regulated protein 94 (GRP94) gene was determined from the isolation and characterization of a genomic clone. Like the mammalian and plant GRP94s, the L. infantum GRP94 sequence possesses both an N-terminal signal peptide and a putative endoplasmic reticulum retention signal, consisting of the C-terminal tetrapeptide EDDL. Thus, L. infantum is the first protozoan organism in which GRP94 has been identified. Southern blot analysis has indicated that this protein is encoded by a single-copy gene. The L. infantum GRP94 gene was expressed in Escherichia coli and the recombinant protein used to evaluate its antigenicity and immunogenicity. Eighty-four percent of sera from dogs with visceral leishmaniasis reacted with the protein, indicating that GRP94 is a potent immunogen during Leishmania infection. Given the immunogenic and antigenic properties shown by the L. infantum GRP94, we think that this protein constitutes a valuable molecule for diagnostic purposes and a potential candidate for studies of protective immunogenicity.
authors.affiliation: Centro de Biología Molecular Severo Ochoa,, Universidad Autónoma de Madrid, 28049 Madrid, Spain.
authors.forename: R
authors.initials: R
authors.lastname: Larreta
authors.forename: M
authors.initials: M
authors.lastname: Soto
authors.forename: C
authors.initials: C
authors.lastname: Alonso
authors.forename: J M
authors.initials: JM
authors.lastname: Requena
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000913:Antibodies, Protozoan / Q000096:biosynthesis; D000953:Antigens, Protozoan / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D003001:Cloning, Molecular; D004285:Dogs; D005260:Female; D015870:Gene Expression; D018840:HSP70 Heat-Shock Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D006801:Humans; D007106:Immune Sera / Q000276:immunology; D007114:Immunization; D018314:Leishmania infantum / Q000235:genetics* / Q000276:immunology; D008565:Membrane Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D051379:Mice; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D016415:Sequence Alignment
chemical_list: D000913:Antibodies, Protozoan; D000953:Antigens, Protozoan; D018840:HSP70 Heat-Shock Proteins; D007106:Immune Sera; D008565:Membrane Proteins; D011994:Recombinant Proteins; C018551:glucose-regulated proteins