Leishmania (Viannia) braziliensis nucleoside triphosphate diphosphohydrolase (NTPDase 1): Localization and in vitro inhibition of promastigotes growth by polyclonal antibodies

• Published in: Experimental parasitology 2012-10, Vol.132 (2), p.293-299
• Main Authors: Porcino, Gabriane Nascimento, Carvalho-Campos, Cristiane, Maia, Ana Carolina Ribeiro Gomes, Detoni, Michelle Lima, Faria-Pinto, Priscila, Coimbra, Elaine Soares, Marques, Marcos José, Juliano, Maria Aparecida, Juliano, Luiz, Diniz, Vanessa Álvaro, Corte-Real, Suzana, Vasconcelos, Eveline Gomes
• Format: Article
• Language: English
• Subjects: adenosinetriphosphatase; Animals; Antibodies, Immobilized - immunology; Antibodies, Protozoan - immunology; Antigens, CD - analysis; Antigens, CD - immunology; antiserum; Apyrase; Apyrase - analysis; Apyrase - antagonists & inhibitors; Apyrase - immunology; ATP diphosphohydrolase; Blotting, Western; cytotoxicity; disease control; Immunocytochemical; Immunoprecipitation; Indexing in process; Isoenzymes - analysis; Isoenzymes - immunology; Leishmania braziliensis; Leishmania braziliensis - enzymology; Leishmania braziliensis - growth & development; Leishmania braziliensis - immunology; Leishmaniasis; Mice; Mice, Inbred BALB C; microscopy; Microscopy, Immunoelectron; mitochondria; NTPase; nucleosides; parasites; parasitology; Peptide; polyclonal antibodies; promastigotes; Rabbits; synthetic peptides
• ISSN: 0014-4894; 1090-2449
• DOI: 10.1016/j.exppara.2012.08.009

[Display omitted] ► Antigenic domain was identified within NTPDase 1 from Leishmania (Viannia) braziliensis. ► Anti-peptides antibodies were used as tools for studies of this isoform. ► It was immunolocalized on the surface and subcellular sites. ► The NTPDase 1 activity was inhibited by immune sera anti-peptides. ► Immune sera anti-peptides significantly reduce in vitro promastigotes growth. Nucleoside triphosphate diphosphohydrolase (NTPDase) activity was recently characterized in Leishmania (Viannia) braziliensis promastigotes (Lb), and an antigenic conserved domain (r82–121) from the specific NTPDase 1 isoform was identified. In this work, mouse polyclonal antibodies produced against two synthetic peptides derived from this domain (LbB1LJ, r82–103; LbB2LJ, r102–121) were used. The anti-LbB1LJ or anti-LbB2LJ antibodies were immobilized on protein A-sepharose and immunoprecipitated the NTPDase 1 of 48kDa and depleted approximately 40% of the phosphohydrolytic activity from detergent-homogenized Lb preparation. Ultrastructural immunocytochemical microscopy identified the NTPDase 1 on the parasite surface and in its subcellular cytoplasmic vesicles, mitochondria, kinetoplast and nucleus. The ATPase and ADPase activities of detergent-homogenized Lb preparation were partially inhibited by anti-LbB1LJ antibody (43–79%), which was more effective than that inhibition (18–47%) by anti-LbB2LJ antibody. In addition, the immune serum anti-LbB1LJ (67%) or anti-LbB2LJ (33%) was cytotoxic, significantly reducing the promastigotes growth in vitro. The results appoint the conserved domain from the L. braziliensis NTPDase as an important target for inhibitor design and the potential application of these biomolecules in experimental protocols of disease control.