Comparative hydrolysis of P2 receptor agonists by NTPDases 1, 2, 3 and 8

Abstract

Nucleoside triphosphate diphosphohydrolases 1, 2, 3 and 8 (NTPDases 1, 2, 3 and 8) are the dominant ectonucleotidases and thereby expected to play important roles in nucleotide signaling. Distinct biochemical characteristics of individual NTPDases should allow them to regulate P2 receptor activation differentially. Therefore, the biochemical and kinetic properties of these enzymes were compared. NTPDases 1, 2, 3 and 8 efficiently hydrolyzed ATP and UTP with Km values in the micromolar range, indicating that they should terminate the effects exerted by these nucleotide agonists at P2X1- and P2Y2,4,11 receptors. Since NTPDase1 does not allow accumulation of ADP, it should terminate the activation of P2Y1,12,13 receptors far more efficiently than the other NTPDases. In contrast, NTPDases 2, 3 and 8 are expected to promote the activation of ADP specific receptors, because in the presence of ATP they produce a sustained (NTPDase2) or transient (NTPDases 3 and 8) accumulation of ADP. Interestingly, all plasma membrane NTPDases dephosphorylate UTP with a significant accumulation of UDP, favoring P2Y6 receptor activation. NTPDases differ in divalent cation and pH dependence, although all are active in the pH range of 7.0-.5. Various NTPDases may also distinctly affect formation of extracellular adenosine and therefore adenosine receptor-mediated responses, since they generate different amounts of the substrate (AMP) and inhibitor (ADP) of ecto-5-nucleotidase, the rate limiting enzyme in the production of adenosine. Taken together, these data indicate that plasma membrane NTPDases hydrolyze nucleotides in a distinctive manner and may therefore differentially regulate P2 and adenosine receptor signaling.

Hyperglycemia alters E-NTPDases, ecto-5′-nucleotidase, and ectosolic and cytosolic adenosine deaminase activities and expression from encephala of adult zebrafish (Danio rerio)

“…While we do not have specific information about the catalytic properties of ecto-nucleotidase isoforms expressed in zebrafish, assuming data from mammals and amphibian, the decrease in ATP hydrolysis seems here could involve NTPDase 2, due to hydrolysis ratio 30:1 (ATP:ADP) that contributes more effectively to ATP hydrolysis when compared with other NTPDases [15,49,50]. In fact, the expression of E-NTPDase 2 isoforms (entpd2a.1 and entpd2a) was significantly decreased in glucose-treated zebrafish.…”

Ectonucleotidases in the kidney

“…Although ATP is dephosphorylated one phosphate at a time, the processing step is favoured, leading to the rapid generation of AMP with little ADP accumulation in the milieu [9][10][11]. Interestingly, and of possible physiological importance, the kinetics are different when UTP is used as substrate, as UDP accumulates transiently, being completely hydrolyzed only after the concentration of UTP has decreased [10]. NTPDase2 has a much greater preference for the hydrolysis of triphosphonucleosides (e.g., ATP) than the diphosphate derivatives (e.g., ADP), and therefore causes accumulation of the latter; NTPDase3 and NTPDase8 also have a preference for the Major pathways are shown in black, others in grey.…”

“…For most of these enzymes, nucleotides derived from other bases (UTP, GTP, TTP, CTP, UDP, GDP, TDP and CDP) can also act as substrates hydrolysis of tri-over diphosphonucleosides, though in these cases the preference is less marked [8,12]. Thus, NTPDases 3 and 8 generate only a transient accumulation of diphosphonucleosides [10,13,14].…”

Characterization of an alternative splice variant of human nucleoside triphosphate diphosphohydrolase 3 (NTPDase3): A possible modulator of nucleotidase activity and purinergic signaling

“…NTPDase5 and 6 have a unique topology in that they have just one membrane-spanning domain and, after cleavage of these respective N-terminal signal peptides, they can be released into the extracellular space as soluble enzymes [4][5][6][7][8][9]. NTPDases that are found at the cell surface modulate signaling mediated by cell-surface purinergic receptors, which are known to control many physiological processes, including blood clotting, pain perception, and smooth muscle contraction [1,10].…”

“…NTPDase5 and 6 greatly prefer diphosphates over triphosphates as substrates [4][5][6][7][8][9]. These differential nucleotide hydrolysis profiles are important for distinguishing the functions of these various NTPDases [10]. Despite these differences in substrate specificity, all family members are characterized by the presence of seven highly conserved domains, called apyrase conserved regions (ACRs [11,12]) and by their dependence upon divalent cations (Mg 2+ or Ca 2+ ) for activity.…”