title: Human and murine immune responses to a novel Leishmania major recombinant protein encoded by members of a multicopy gene family.
abstract: Vaccination of BALB/c mice with Leishmania major promastigote culture filtrate proteins plus Corynebacterium parvum confers resistance to infection with L. major. To define immunogenic components of this protein mixture, we used sera from vaccinated mice to screen an L. major amastigote cDNA expression library. One of the immunoreactive clones thus obtained encoded a novel protein of L. major with a molecular mass of 22.1 kDa. The predicted amino acid sequence of this clone exhibited significant homology to eukaryotic thiol-specific-antioxidant (TSA) proteins. Therefore, we have designated this protein L. major TSA protein. Southern blot hybridization analyses indicate that there are multiple copies of the TSA gene in all species of Leishmania analyzed. Northern blot analyses demonstrated that the TSA gene is constitutively expressed in L. major promastigotes and amastigotes. Recombinant TSA protein containing an amino-terminal six-histidine tag was expressed in Escherichia coli with the pET17b system and was purified to homogeneity by affinity chromatography. Immunization of BALB/c mice with recombinant TSA protein resulted in the development of strong cellular immune responses and conferred protective immune responses against infection with L. major when the protein was combined with interleukin 12. In addition, recombinant TSA protein elicited in vitro proliferative responses from peripheral blood mononuclear cells of human leishmaniasis patients and significant TSA protein-specific antibody titers were detected in sera of both cutaneous-leishmaniasis and visceral-leishmaniasis patients. Together, these data suggest that the TSA protein may be useful as a component of a subunit vaccine against leishmaniasis.
authors.affiliation: Infectious Disease Research Institute, University of Washington, Seattle, Washington, USA.
authors.forename: J R
authors.initials: JR
authors.lastname: Webb
authors.forename: A
authors.initials: A
authors.lastname: Campos-Neto
authors.forename: P J
authors.initials: PJ
authors.lastname: Ovendale
authors.forename: T I
authors.initials: TI
authors.lastname: Martin
authors.forename: E J
authors.initials: EJ
authors.lastname: Stromberg
authors.forename: R
authors.initials: R
authors.lastname: Badaro
authors.forename: S G
authors.initials: SG
authors.lastname: Reed
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D005260:Female; D006801:Humans; D015151:Immunoblotting; D018664:Interleukin-12 / Q000494:pharmacology; D018320:Leishmania major / Q000276:immunology*; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D005810:Multigene Family; D015800:Protozoan Proteins / Q000235:genetics / Q000276:immunology*; D016052:Protozoan Vaccines / Q000276:immunology; D011994:Recombinant Proteins / Q000276:immunology
chemical_list: D015800:Protozoan Proteins; D016052:Protozoan Vaccines; D011994:Recombinant Proteins; D018664:Interleukin-12