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Characterization of ectonucleoside triphosphate diphosphohydrolase ( E‐NTPDase ; EC 3.6.1.5) activity in mouse peritoneal cavity cells
Author(s) -
Dias Dhébora Albuquerque,
Penteado Bruna,
Santos Lucas Derbocio,
Santos Pedro Mendes,
Arruda Carla Cardozo Pinto,
Schetinger Maria Rosa Chitolina,
Leal Daniela Bitencourt Rosa,
Jaques Jeandre Augusto
Publication year - 2017
Publication title -
cell biochemistry and function
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.933
H-Index - 61
eISSN - 1099-0844
pISSN - 0263-6484
DOI - 10.1002/cbf.3281
Subject(s) - atp hydrolysis , adenosine triphosphate , sodium azide , nucleotide , atpase , adenosine diphosphate , chemistry , enzyme , sodium fluoride , biochemistry , hydrolysis , sodium , nucleoside triphosphate , suramin , adenosine , adenosine monophosphate , peritoneal cavity , adenine nucleotide , microbiology and biotechnology , biology , platelet , in vitro , fluoride , inorganic chemistry , platelet aggregation , organic chemistry , anatomy , immunology , gene
This study aimed to characterize the activity of ectonucleoside triphosphate diphosphohydrolase (E‐NTPDase; EC 3.6.1.5) in peritoneal cavity cells from BALB/c mice. E‐NTPDase was activated in the presence of both calcium (1.5mM) and magnesium (1.5mM) ions. However, the activity was higher in the presence of Ca 2+ . A pH of 8.5 and temperature of 37°C were the optimum conditions for catalysis. The apparent Km values were 0.51mM and 0.66mM for the hydrolysis of adenosine triphosphate (ATP) and adenosine diphosphate (ADP), respectively. The Vmax values were 136.4 and 120.8 nmol Pi/min/mg of protein for ATPase and ADPase activity, respectively. Nucleotide hydrolysis was inhibited in the presence of sodium azide (20mM, ATP: P < .05; ADP: P < .001), sodium fluoride (20mM; ATP and ADP: P < .001), and suramin (0.3mM; ATP: P < .01; ADP: P < .05), which is a known profile for NTPDase inhibition. Although all of the diphosphate and triphosphate nucleotides that were tested were hydrolyzed, enzyme activity was increased when adenine nucleotides were used as substrates. Finally, we stress that knowledge of the E‐NTPDase catalytic biochemical properties in mouse peritoneal cavity cells is indispensable for properly determining its activity, as well as to fully understand the immune response profile in both healthy and sick cells.