title: Leishmania amazonensis: characterization of an ecto-3'-nucleotidase activity and its possible role in virulence.
abstract: Ecto-3'-nucleotidase/nuclease (3'NT/NU) is a membrane-bound enzyme that plays a key role in the nutrition of Leishmania sp. protozoan parasites. This enzyme generates nucleosides via hydrolyzes of 3'mononucleotides and nucleic acids, which enter the cell by specific transporters. In this work, we identify and characterize Leishmania amazonensis ecto-3'-nucleotidase activity (La3'-nucleotidase), report ammonium tetrathiomolybdate (TTM) as a novel La3'-nucleotidase inhibitor and approach the possible involvement of ecto-3'-nucleotidase in cellular adhesion. La3'-nucleotidase presented characteristics similar to those reported for the class I single-strand nuclease family; a molecular weight of approximately 40 kDa and optimum activity in an alkaline pH range were observed. Although it is conserved among the genus, La3'-nucleotidase displays different kinetic properties; it can be inhibited by vanadate, molybdate and Cu(2+) ions. Interestingly, ecto-3'-nucleotidase activity is 60-fold higher than that of ecto-5'-nucleotidase in L. amazonensis. Additionally, ecto-3'-nucleotidase activity is two-fold higher in virulent L. amazonensis cells than in avirulent ones. Notably, macrophage-parasite attachment/invasion was increased by 400% in the presence of adenosine 3'-monophosphate (3'AMP); however, this effect was reverted by TTM treatment. We believe that La3'-nucleotidase may play a significant role in the generation of adenosine, which may contribute to mammalian host immune response impairment and establishment of infection.
authors.affiliation: Instituto de Bioquímica Médica, Universidade Federal do Rio de Janeiro, CCS, Bloco H, Cidade Universitária, Ilha do Fundão, 21941-590 Rio de Janeiro, RJ, Brazil.
authors.forename: Rafael
authors.initials: R
authors.lastname: Paletta-Silva
authors.forename: Danielle P
authors.initials: DP
authors.lastname: Vieira
authors.forename: Rodrigo
authors.initials: R
authors.lastname: Vieira-Bernardo
authors.forename: David
authors.initials: D
authors.lastname: Majerowicz
authors.forename: Katia C
authors.initials: KC
authors.lastname: Gondim
authors.forename: Marcos André
authors.initials: MA
authors.lastname: Vannier-Santos
authors.forename: Angela H
authors.initials: AH
authors.lastname: Lopes
authors.forename: José Roberto
authors.initials: JR
authors.lastname: Meyer-Fernandes
mesh_terms: D000249:Adenosine Monophosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D006224:Cricetinae; D005260:Female; D006790:Host-Parasite Interactions; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007894:Leishmania mexicana / Q000145:classification / Q000201:enzymology* / Q000472:pathogenicity*; D017737:Macrophages, Peritoneal / Q000469:parasitology*; D051379:Mice; D008807:Mice, Inbred BALB C; D009708:Nucleotidases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010802:Phylogeny; D016415:Sequence Alignment; D014774:Virulence
chemical_list: D000249:Adenosine Monophosphate; D009708:Nucleotidases; C040470:3'-nucleotidase