title: Characterization of NTPDase (NTPDase1; ecto-apyrase; ecto-diphosphohydrolase; CD39; EC 3.6.1.5) activity in human lymphocytes.
abstract: Human lymphocytes contain NTPDase (NTPDase-1; ecto-apyrase; ecto-diphosphohydrolase; CD39; EC 3.6.1.5), a cation-dependent enzyme that hydrolyzes ATP and ADP and also other di- and triphosphate nucleosides, acting at an optimum pH of 8.0. A significant inhibition of ATP and ADP hydrolysis (P<0.05) was observed in the presence of 20 mM sodium azide. NTPDase inhibitors, 20 mM sodium fluoride, 0.2 mM trifluoperazine and 0.3 mM suramin, significantly decreased ATP and ADP hydrolysis (P<0.05) and ADP hydrolysis was only inhibited by 0.5 mM orthovanadate (P<0.05). ATP and ADP hydrolysis was not inhibited in the presence of 0.01 mM Ap5A (P1,P5-di(adenosine-5')pentaphosphate), 0.1 mM ouabain, 1 mM levamisole, 2 microg/mL oligomycin, 0.1 mM N-ethylmaleimide (NEM), or 5 mM sodium azide. With respect to kinetic behavior, apparent K(m) values of 77.6+/-10.2 and 106.8+/-21.0 microM, and V(max) values of 68.9+/-8.1 and 99.4+/-8.5 (mean+/-S.E., n=3) nmol Pi/min/mg protein were obtained for ATP and ADP, respectively. A Chevilard plot demonstrated that only one enzymatic site is responsible for the hydrolysis of ATP and ADP. The presence of CD39 was determined by flow cytometry, showing a low density of 2.72+/-0.24% (mean+/-S.E.; n=30) in human peripheral lymphocytes. The study of NTPDase activity in human lymphocytes may be important to determine the immune response status against infectious agents related to ATP and ADP hydrolysis.
authors.affiliation: Departamento de Bioquímica, Instituto de Ciências Básicas da Saúde, Universidade Federal do Rio Grande do Sul, Rua Ramiro Barcellos, 2600-Anexo, 90035-003. Porto Alegre, RS, Brazil.
authors.forename: Daniela B R
authors.initials: DB
authors.lastname: Leal
authors.forename: Cristiane A
authors.initials: CA
authors.lastname: Streher
authors.forename: Tiago N
authors.initials: TN
authors.lastname: Neu
authors.forename: Fábio P
authors.initials: FP
authors.lastname: Bittencourt
authors.forename: Cláudio A M
authors.initials: CA
authors.lastname: Leal
authors.forename: José E P
authors.initials: JE
authors.lastname: da Silva
authors.forename: Vera M
authors.initials: VM
authors.lastname: Morsch
authors.forename: Maria R C
authors.initials: MR
authors.lastname: Schetinger
mesh_terms: D000244:Adenosine Diphosphate / Q000378:metabolism; D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism; D015703:Antigens, CD / Q000378:metabolism*; D001081:Apyrase / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008214:Lymphocytes / Q000201:enzymology*; D013379:Substrate Specificity
chemical_list: D015703:Antigens, CD; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; D001081:Apyrase; C067869:CD39 antigen