Giardia duodenalis: biochemical characterization of an ecto-5'-nucleotidase activity.
In this work, we biochemically characterized the ecto-5'-nucleotidase activity present on the surface of the living trophozoites of Giardia duodenalis. Two sequences of the 5'-nucleotidase family protein were identified in the Giardia genome. Anti-mouse CD73 showed a high reaction with the cell surface of parasites. At pH 7.2, intact cells were able to hydrolyze 5'-AMP at a rate of 10.66 ± 0.92 nmol Pi/h/10(7) cells. AMP is the best substrate for this enzyme, and the optimum pH lies in the acidic range. No divalent cations had an effect on the ecto-5'-nucleotidase activity, and the same was seen for NaF, an acid phosphatase inhibitor. Ammonium molybdate, a potent inhibitor of nucleotidases, inhibited the enzyme activity in a dose-dependent manner. The presence of adenosine in the culture medium negatively modulated the enzyme. The results indicate the existence of an ecto-5'-nucleotidase that could play a role in the salvage of purines.
Russo-Abrahão
Thais
T
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.
Cosentino-Gomes
Daniela
D
Daflon-Yunes
Nathalia
N
Meyer-Fernandes
José Roberto
JR
D015720:5'-Nucleotidase / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D000227:Adenine Nucleotides / Q000378:metabolism*; D000241:Adenosine / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D004305:Dose-Response Relationship, Drug; D005434:Flow Cytometry; D005872:Giardia / Q000201:enzymology* / Q000235:genetics / Q000276:immunology; D006863:Hydrogen-Ion Concentration; D051379:Mice; D008969:Molecular Sequence Data; D008982:Molybdenum / Q000494:pharmacology; D016415:Sequence Alignment; D013379:Substrate Specificity
D000227:Adenine Nucleotides; D008982:Molybdenum; D015720:5'-Nucleotidase; D000241:Adenosine; C022175:ammonium molybdate