Kinetic and biochemical characterization of an ecto-nucleotide pyrophosphatase/phosphodiesterase (EC 3.1.4.1) in cells cultured from submandibular salivary glands of rats.
The participation of ecto-nucleotide pyrophosphatase/phosphodiesterase (E-NPP) activity in the nucleotide hydrolysis by salivary gland cells of rats was evaluated using p-nitrophenyl 5'-thymidine monophosphate (p-Nph-5'-TMP) as a substrate for this enzyme. We investigated the biochemical characteristics of this ectoenzyme in cells cultured from submandibular salivary glands of rats. Primary cell cultures demonstrated ecto-nucleotide pyrophosphatase/phosphodiesterase (E-NPP) activities, which could be observed by extracellular hydrolysis of p-Nph-5'-TMP and other biochemical characteristics such as dependence of metal ions, dependence of pH alkaline and inactivation by a metal ion chelator. The Km value for the hydrolysis of p-Nph-5'-TMP was 280.7+/-34.2 microM (mean+/-S.D., n=4) and Vmax was 721.31+/-225nmol p-nitrophenol/min/mg (mean+/-S.D., n=4). We suggest that E-NPP is co-localized with an ecto-ATP diphosphohydrolase/ecto-NTPDase and an ecto-5'-nucleotidase, since these enzymes probably act under different conditions. It may be postulated that the physiological role for these ecto-enzymes is to terminate the action of the co-transmitter ATP, generating adenosine.
Henz
Sandra Liana
SL
Departamento de Bioquímica, Instituto de Ciências Básicas da Saúde, Universidade Federal do Rio Grande do Sul, Rua Ramiro Barcelos 2600-Anexo,90035-003, Porto Alegre, RS, Brazil.
Fürstenau
Cristina Ribas
CR
Chiarelli
Rafael Augusto
RA
Sarkis
João José Freitas
JJ
D000244:Adenosine Diphosphate / Q000737:chemistry / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000737:chemistry / Q000378:metabolism*; D000704:Analysis of Variance; D000818:Animals; D002478:Cells, Cultured; D004789:Enzyme Activation; D006868:Hydrolysis; D009711:Nucleotides / Q000737:chemistry / Q000378:metabolism*; D010727:Phosphoric Diester Hydrolases / Q000737:chemistry / Q000378:metabolism* / Q000493:pharmacokinetics; D011755:Pyrophosphatases / Q000737:chemistry / Q000378:metabolism* / Q000493:pharmacokinetics; D051381:Rats; D013363:Submandibular Gland / Q000201:enzymology* / Q000378:metabolism
D009711:Nucleotides; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D010727:Phosphoric Diester Hydrolases; C065962:ectonucleotide pyrophosphatase phosphodiesterase 1; D011755:Pyrophosphatases