title: An antigenic domain within a catalytically active Leishmania infantum nucleoside triphosphate diphosphohydrolase (NTPDase 1) is a target of inhibitory antibodies.
abstract: We identified a shared B domain within nucleoside triphosphate diphosphohydrolases (NTPDases) of plants and parasites. Now, an NTPDase activity not affected by inhibitors of adenylate kinase and ATPases was detected in Leishmania infantum promastigotes. By non-denaturing gel electrophoresis of detergent-homogenized promastigote preparation, an active band hydrolyzing nucleosides di- and triphosphate was visualized and, following SDS-PAGE and silver staining was identified as a single polypeptide of 50kDa. By Western blots, it was recognized by immune sera raised against potato apyrase (SA), r-pot B domain (SB), a recombinant polypeptide derived from the potato apyrase, and LbB1LJ (SC) or LbB2LJ (SD), synthetic peptides derived from the Leishmania NTPDase 1, and by serum samples from dogs with visceral leishmaniasis, identifying the antigenic L. infantum NTPDase 1 and, also, its conserved B domain (r83-122). By immunoprecipitation assays and Western blots, immune sera SA and SB identified the catalytically active NTPDase 1 in promastigote preparation. In addition, the immune sera SB (44%) and SC or SD (87-99%) inhibited its activity, suggesting a direct effect on the B domain. By ELISA, 37%, 45% or 50% of 38 infected dogs were seropositive for r-pot B domain, LbB1LJ and LbB2LJ, respectively, confirming the B domain antigenicity.
authors.affiliation: Departamento de Bioquímica, Laboratório de Estrutura e Função de Proteínas, Pós-Graduação em Imunologia e DIP/Genética e Biotecnologia, Instituto de Ciências Biológicas, Universidade Federal de Juiz de Fora, Juiz de Fora, MG, Brazil.
authors.forename: Ana Carolina Ribeiro Gomes
authors.initials: AC
authors.lastname: Maia
authors.forename: Gabriane Nascimento
authors.initials: GN
authors.lastname: Porcino
authors.forename: Michelle de Lima
authors.initials: Mde L
authors.lastname: Detoni
authors.forename: Nayara Braga
authors.initials: NB
authors.lastname: Emídio
authors.forename: Danielle Gomes
authors.initials: DG
authors.lastname: Marconato
authors.forename: Priscila
authors.initials: P
authors.lastname: Faria-Pinto
authors.forename: Melissa Regina
authors.initials: MR
authors.lastname: Fessel
authors.forename: Alexandre Barbosa
authors.initials: AB
authors.lastname: Reis
authors.forename: Luiz
authors.initials: L
authors.lastname: Juliano
authors.forename: Maria Aparecida
authors.initials: MA
authors.lastname: Juliano
authors.forename: Marcos José
authors.initials: MJ
authors.lastname: Marques
authors.forename: Eveline Gomes
authors.initials: EG
authors.lastname: Vasconcelos
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D015703:Antigens, CD / Q000737:chemistry* / Q000276:immunology* / Q000302:isolation & purification / Q000378:metabolism; D000953:Antigens, Protozoan / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D001081:Apyrase / Q000737:chemistry* / Q000276:immunology* / Q000302:isolation & purification / Q000378:metabolism; D004285:Dogs; D018314:Leishmania infantum / Q000201:enzymology* / Q000235:genetics / Q000276:immunology*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D016415:Sequence Alignment
chemical_list: D015703:Antigens, CD; D000953:Antigens, Protozoan; D001081:Apyrase; C067869:CD39 antigen