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At least 19 recordsLinked to original sources
PubMed1997

Immunolocalization of the ecto-ATPase and ecto-apyrase in chicken gizzard and stomach. Purification and N-terminal sequence of the stomach ecto-apyrase.

We have examined the in vivo localization of extracellular ecto-ATPase and ecto-apyrase (ATPDase) in adult chicken gizzard and stomach by immunofluorescence and laser scanning confocal microscopy. In chicken gizzard, the ecto-ATPase was distributed in discrete clusters restricted to the sarcolemma of the smooth muscle cells. Anti-ecto-apyrase antibody detected a single 80-kDa band (putative apyrase) in Western blots of both chicken gizzard membrane extracts and partially purified anion exchange fractions, but the antibody did not detect ecto-apyrase in immunolabeled gizzard cryosections. In adult chicken stomach, the ecto-apyrase was observed at the apical membrane of the glandular oxyntico-peptic cells as described in previous immunoperoxidase studies (Stout, J. G., R. S. Strobel, and T. L. Kirley (1995) Biochem. Mol. Biol. Int. 36, 529-535). However, ecto-ATPase was clustered in the sarcolemma of the organized layer of circular smooth muscle and in smooth muscle cells of the septa surrounding the glandular tissue, but not in the glandular cells containing the ecto-apyrase. The findings indicate compartmentalization of the two related extracellular nucleotide hydrolyzing enzymes and suggest differential functions that are specialized for different regions of the chicken stomach. We also partially purified the ecto-apyrase of chicken stomach, an 80-kDa membrane glycoprotein. Chicken stomach membranes were solubilized in digitonin, glycoproteins were separated from solubilized proteins by lectin chromatography, and nucleotide-binding glycoproteins were selected by immobilized Cibacron blue chromatography. Further purification by size exclusion and anion exchange chromatography yielded purification of 94-fold. The ATPase specific activity of the purified stomach ecto-apyrase was 75,000 micromol of Pi/mg of protein/h, and the purified preparation consisted of a major band (55% of total protein) at 80 kDa. The purified enzyme could be deglycosylated with peptide N-glycosidase-F to a core molecular mass of 54 kDa. The N-terminal sequence of the 80-kDa stomach ecto-apyrase band (which reacted with anti-ecto-ATPDase antibodies) was determined to be: MEYKGKVVAGLLTATWV. Immunological cross-reactivity data indicate that the stomach 80-kDa protein isolated is an ecto-apyrase and is related to both the chicken liver and oviduct ecto-ATPDase enzymes characterized earlier, as well as to the human lymphoid cell activation antigen, CD39.

Adenosine Triphosphatases↗
PubMed1989

Comparative subcellular distribution of apyrase from animal and plant sources. Characterization of microsomal apyrase.

1. Apyrase (ATP: diphosphohydrolase) has been found in the microsomal fraction of rat salivary gland, mammary gland and uterus. 2. This enzyme, already described in plant tissue, is mainly present as a soluble polypeptide in tubers of Solanum tuberosum. 3. A fraction of this enzyme is associated with the microsomal fraction with a higher specific activity than the soluble one, for either ATP or ADP as substrate. 4. Apyrase bound to microsomes from rat and potato tissues was characterized in its substrate specificity and effect of inhibitors. 5. The Km values for ATP and ADP, optimum pH and metal ion requirement were determined. 6. A characteristic common to the microsomal and soluble apyrases is the stimulatory effect of a potato activator protein of soluble plant apyrase. 7. The microsomal-bound apyrase from rat and potato tissues were solubilized and subjected to size-exclusion chromatography. 8. The mammary gland and salivary gland apyrases eluted as molecular aggregates, in contrast to the uterus and potato enzyme.

Animals↗
PubMed1997

8-Azido-nucleotides as substrates of Torpedo electric organ apyrase. effect of photoactivation on apyrase activity.

Ecto-apyrase is a widespread enzymatic activity that hydrolyses tri- and diphosphonucleotides and consequently controls the amount of available extracellular ATP and ADP. In the nervous system, purines have important neuromodulatory actions, acting at pre- and postsynaptic sites, and consequently, ecto-apyrase may play an indirect role in the modulation of nucleotide- and nucleoside-mediated processes. The azido-nucleotides have been largely employed to characterize the nucleotide binding sites of several proteins. In the present work the azido-nucleotides are described as putative substrates for apyrase activity in a presynaptic plasma membrane preparation (PSPM) from the Torpedo electric organ. Both 8-N3-ATP and 8-N3-ADP were hydrolyzed in a calcium-dependent manner showing Vmax of 23.8 +/- 4.8 and 14.5 +/- 3 U/mg of protein, and Km values (in microM) of 116 +/- 39 and 119 +/- 4, respectively. Vmax for calcium-dependent hydrolysis of ATP and ADP were significantly higher: 59.2 +/- 3.9 and 32.9 +/- 3.5 U/mg of protein respectively, while Km values did not show any significant differences regarding azido-nucleotides: 83.8 +/- 12 microM for Ca2+-ATP and 121 +/- 34 microM for Ca2+-ADP. The photoactivation of the PSPM in the presence of the azido-derivatives results in an irreversible inactivation of apyrase activity, showing an IC50 of 10 microM and a maximal inhibitory effect of 38 and 60% on Ca2+-ATPase and Ca2+-ADPase activities. Apyrase was protected from inactivation by nucleotides that are natural substrates for this enzymatic activity and also by AMP while adenosine did not protect from apyrase inhibition.

Adenosine Diphosphate↗
PubMed2001

The salivary apyrase of the blood-sucking sand fly Phlebotomus papatasi belongs to the novel Cimex family of apyrases.

Apyrases are enzymes that hydrolyze nucleotide di- and triphosphates to orthophosphate and mononucleotides. At least two families of enzymes, belonging to the 5'-nucleotidase and to the actin/heat shock 70/sugar kinase superfamily, have evolved independently to serve the apyrase reaction. Both families require either Ca(2+) or Mg(2+) for their action. A novel apyrase enzyme sequence, with no homology to any other known protein sequence, was found recently in the salivary glands of the hematophagous bed bug Cimex lectularius. This enzyme functions exclusively with Ca(2+). Here, we report the finding of a cDNA similar to that of the C. lectularius salivary apyrase isolated from a salivary gland cDNA library of Phlebotomus papatasi. Transfection of insect cells with the P. papatasi salivary gland apyrase cDNA resulted in the secretion of a Ca(2+)-dependent apyrase whose activity was indistinguishable from that in salivary homogenates of P. papatasi. Homologous sequences were found in humans, in another sand fly (Lutzomyia longipalpis), in the fruit fly Drosophila melanogaster, in the nematode Caenorhabditis elegans and in the protozoan Cryptosporidium parvum, indicating that this family of enzymes is widespread among animal species.

Amino Acid Sequence↗
PubMed1997

Characterization of brain ecto-apyrase: evidence for only one ecto-apyrase (CD39) gene.

A rat brain cDNA coding for ecto-(Ca,Mg)-apyrase activity was isolated using human CD39 cDNA and functionally expressed in COS-7 cells. The gene codes for a protein with high similarity to human (75% identity) and murine (90% identity) CD39. It is expressed in primary neurons and astrocytes in cell culture as well as in kidney, liver, muscle and spleen. Southern analysis of the mouse genome suggests that there may be a single copy of the ecto-apyrase gene. Interestingly, the human CD39 gene cytologically co-localizes with the susceptibility gene involved in human partial epilepsy with audiogenic symptoms; such a coincidence is consistent with reports on the deficiency of ecto-apyrase activity in the brains of humans with temporal lobe epilepsy and in those of mice with audiogenic seizures.

Adenosine Triphosphatases↗
PubMed2000

Regulation of a recombinant pea nuclear apyrase by calmodulin and casein kinase II.

A cDNA encoding a pea nuclear apyrase was previously cloned. Overexpressions of a full-length and a truncated cDNA have been successfully expressed in Escherichia coli BL21(DE3). The resulting fusion proteins, apyrase and the C-terminus (residues 315-453) of apyrase, were used for calmodulin (CaM) binding and phosphorylation studies. Fusion protein apyrase but not the C-terminus of apyrase can be recognized by polyclonal antibody pc480. This suggested that the motif recognized by pc480 was located in the N-terminal region of apyrase. The recombinant apyrase protein also showed an activity 70 times higher than that of endogenous apyrase using ATP as a substrate. The recombinant apyrase has a preference for ATP more than other nucleoside triphosphate substrates. CaM can bind to recombinant apyrase, but not to the C-terminus of apyrase. This implies that the CaM-binding domain must be in the first 315 amino acids of the N-terminal region of apyrase. We found that one segment from residue 293 to 308 was a good candidate for the CaM-binding domain. This segment 293 FNKCKNTIRKALKLNY 308 has a basic amphiphilic-helical structure, which shows the predominance of basic residues on one side and hydrophobic residues on the other when displayed on a helical wheel plot. Using the gel mobility shift binding assay, this synthetic peptide was shown to bind to CaM, indicating that it is the CaM-binding domain. Both recombinant apyrase and the C-terminus of apyrase can be phosphorylated by a recombinant human protein kinase CKII. Phosphorylation does not affect CaM binding to recombinant apyrase. However, CaM does inhibit CKII phosphorylation of recombinant apyrase and this inhibition can be blocked by 5 mM EGTA.

Amino Acid Sequence↗
PubMed1996

Comparative studies on animal and plant apyrases (ATP diphosphohydrolase EC 3.6.1.5) with application of immunological techniques and various ATPase inhibitors.

Apyrase activity has been found in tissue of all investigated plant species. Seedlings soluble fractions accounted for 45-75% of the cell apyrase activity, whereas the apyrases isolated from microsomes accounted for 0.2-7% of the total homogenate activity. The ratio of the rate of ATP hydrolysis to the rate of ADP hydrolysis, Ksh, divides the apyrases into two groups: of Ksh > 1 (enzymes from most of monocot plants and bovine tissues) and of Ksh < 1 (enzymes from dicot plants). Triflupromazine strongly decreased the activity of wheat and bovine apyrases (first group) and does not inhibit the activity of the enzyme from potato (second group). Analysed apyrases reveal a significant antigenic diversity. Antibodies developed against soluble potato apyrase have no affinity to apyrase from microsomes of wheat seedlings. Immunological analysis confirmed that ATPase and ADPase activities of potato apyrase were associated with one protein. Apyrases, including animal ones, are insensitive to ATPases inhibitors and reagents of SH groups, whereas sodium deoxycholate inhibits all of the studied enzymes. NaF decreases activity plant enzymes, whereas erythrosine B and NaN3 only decreases bovine apyrases.

Adenosine Triphosphatases↗
PubMed2004

Enhanced ecto-apyrase activity of stimulated endothelial or mesangial cells is downregulated by glucocorticoids in vitro.

Endothelial as well as mesangial cells show enhanced activity of ecto-apyrase following pro-inflammatory stimulation in vitro. Since this ecto-enzyme appears to be able to regulate plasma hemopexin, which latter molecule plays a role in the pathogenesis of corticosteroid responsive nephrotic syndrome, the question was raised whether glucocorticoids are potentially able to downregulate ecto-apyrase activity of these cells. Therefore, cell cultures of endothelial or mesangial were stimulated with or without lipopolysaccharide (10 ng/ml). Parallel cultures were supplemented with prednisolone with or without the glucocorticoid receptor antagonist mifepristone in various concentrations. After 24 h, cytospins were prepared and cytochemically stained for ecto-apyrase activity. mRNA for apyrase of these cells was detected using reverse transcription-polymerase chain reaction (RT-PCR). Apyrase activity of either cells or soluble apyrase (0.16 U/ml buffer) with or without supplementation of prednisolone were biochemically assayed for their phosphatase activity. The results show significantly decreased ecto-apyrase activity of lipopolysaccharide-stimulated cells after treatment with prednisolone as compared to non-prednisolone-treated cells. Preincubation with mifepristone did not inhibit the effect of prednisolone. Identical mRNA signals for apyrase were found in prednisolone and non-prednisolone-treated cells. Interestingly, soluble apyrase also showed a significant decrease of activity following preincubation with prednisolone. It is concluded that prednisolone is able to downregulate ecto-apyrase of stimulated endothelial or mesangial cells, which may potentially inhibit the conversion of hemopexin to its pro-inflammatory isoform. As blocking of the cytosolic glucocorticoid receptor showed no effect upon the prednisolone action, whereas prednisolone is able to affect soluble apyrase per se, it is felt that this particular action of prednisolone may (at least partly) be mediated through a non-genomic pathway.

Antigens, CD↗
PubMed1998

Purification, cloning, and expression of an apyrase from the bed bug Cimex lectularius. A new type of nucleotide-binding enzyme.

An enzyme that hydrolyzes the phosphodiester bonds of nucleoside tri- and diphosphates, but not monophosphates, thus displaying apyrase (EC 3.6.1.5) activity, was purified from salivary glands of the bed bug, Cimex lectularius. The purified C. lectularius apyrase was an acidic protein with a pI of 5.1 and molecular mass of approximately 40 kDa that inhibited ADP-induced platelet aggregation and hydrolyzed platelet agonist ADP with specific activity of 379 units/mg protein. Amplification of C. lectularius cDNA corresponding to the N-terminal sequence of purified apyrase produced a probe that allowed identification of a 1.3 kilobase pair cDNA clone coding for a protein of 364 amino acid residues, the first 35 of which constituted the signal peptide. The processed form of the protein was predicted to have a molecular mass of 37.5 kDa and pI of 4.95. The identity of the product of the cDNA clone with native C. lectularius apyrase was proved by immunological testing and by expressing the gene in a heterologous host. Immune serum made against a synthetic peptide with sequence corresponding to the C-terminal region of the predicted cDNA clone recognized both C. lectularius apyrase fractions eluted from a molecular sieving high pressure liquid chromatography and the apyrase active band from chromatofocusing gels. Furthermore, transfected COS-7 cells secreted a Ca2+-dependent apyrase with a pI of 5.1 and immunoreactive material detected by the anti-apyrase serum. C. lectularius apyrase has no significant sequence similarity to any other known apyrases, but homologous sequences have been found in the genome of the nematode C. elegans and in mouse and human expressed sequence tags from fetal and tumor EST libraries.

Amino Acid Sequence↗
PubMed1996

Apyrase administration prolongs discordant xenograft survival.

Platelet thrombi and vascular inflammation are prominent features of discordant xenograft rejection. The purinergic nucleotides ATP and ADP, which are secreted from platelets and released by injured endothelial cells (EC), are important mediators of these reactions. Quiescent EC express the ectoenzyme ATP-diphosphohydrolase (ATPDase; an apyrase), which exerts an important thromboregulatory function by hydrolyzing both ATP and ADP. We have shown that ATPDase activity is rapidly lost from the surface of the EC following ischemia-reperfusion injury and during xenograft rejection. The aim of this study was to supplement ATPDase activity within xenografts by infusion of soluble apyrases, and thereby validate the importance of local ATPDase activity in the modulation of xenograft rejection. Lewis rats underwent heterotopic cardiac xenografting from guinea pigs and apyrase was administered intravenously (200 U/kg) as a single dose to evaluate effects on hyperacute rejection (HAR). This initial dose was followed by a continuous apyrase infusion (8.0 U/kg/hr) directly into the graft aorta in combination with systemic cobra venom factor (CVF) administration to deplete complement when delayed xenograft rejection (DXR) was studied. Functional apyrase levels in vivo were assessed by the capacity of blood samples taken at the time of surgery and rejection to inhibit platelet aggregation in vitro. Apyrase administration significantly prolonged graft survival in HAR and DXR. Functional assays showed inhibition of platelet aggregation suggesting effective systemic antiaggregatory effects of the administered apyrases. Histologic studies showed that apyrase administration abrogated local platelet aggregation and activation in HAR and DXR. Our data demonstrate that local administration of apyrase prolonged discordant xenograft survival. These observations emphasize the potential importance of purinergic mediators in platelet activation during xenograft rejection.

Adenosine Diphosphate↗
PubMed1999

Glycosylation is essential for functional expression of a human brain ecto-apyrase.

The importance of N-linked glycosylation for the function and oligomerization of an E-type ATPase was examined by using tunicamycin and peptide N-glycosidase F, two agents used to prevent and remove glycosylations, respectively. The cDNA encoding a human ecto-apyrase (HB6), predicted to have seven N-linked glycosylation sites, was transiently expressed in mammalian COS cells and the resulting membrane preparations were treated with peptide N-glycosidase F (PNGase-F). PNGase-F caused a decrease in the apparent molecular weight of the protein (consistent with glycan removal) and a decrease in enzymatic activity over time. The ecto-apyrase was also expressed in the presence of tunicamycin, which completely prevented N-linked glycosylation, resulting in a nonglycosylated core protein devoid of ATP and ADP hydrolyzing activity. However, control and tunicamycin-treated cells expressed the enzyme to similar levels and localization. Interestingly, the quaternary structure of this E-type ATPase appears to be dependent upon the presence of glycan chains. The glycosylated ecto-apyrase exists as a homodimer in situ as assessed by both size-exclusion chromatography of detergent-solubilized ecto-apyrase and cross-linking of membrane-bound ecto-apyrase, in contrast to the enzymatically deglycosylated ecto-apyrase and the tunicamycin-treated ecto-apyrase. These results suggest that glycosylation is necessary for homooligomerization and nucleotide hydrolyzing activity, but not for expression and plasma membrane localization of the E-type ATPase. Similar results were obtained with another human ecto-apyrase, CD39, suggesting that the importance of glycosylation may be generalized to all membrane-bound E-type ATPases.

Adenosine Triphosphatases↗
PubMed2004

Triatoma infestans apyrases belong to the 5'-nucleotidase family.

Apyrases are nucleoside triphosphate-diphosphohydrolases (EC 3.6.1.5) present in a variety of organisms. The apyrase activity found in the saliva of hematophagous insects is correlated with the prevention of ADP-induced platelet aggregation of the host during blood sucking. Purification of apyrase activity from the saliva of the triatomine bug Triatoma infestans was achieved by affinity chromatography on oligo(dT)-cellulose and gel filtration chromatography. The isolated fraction includes five N-glycosylated polypeptides of 88, 82, 79, 68 and 67 kDa apparent molecular masses. The isolated apyrase mixture completely inhibited aggregation of human blood platelets. Labeling with the ATP substrate analogue 5'-p-fluorosulfonylbenzoyladenosine showed that the five species have ATP-binding characteristic of functional apyrases. Furthermore, tandem mass spectroscopy peptide sequencing showed that the five species share sequence similarities with the apyrase from Aedes aegypti and with 5'-nucleotidases from other species. The complete cDNA of the 79-kDa enzyme was cloned, and its sequence confirmed that it encodes for an apyrase belonging to the 5'-nucleotidase family. The gene multiplication leading to the unusual salivary apyrase diversity in T. infestans could represent an important mechanism amplifying the enzyme expression during the insect evolution to hematophagy, in addition to an escape from the host immune response, thus enhancing acquisition of a meal by this triatomine vector of Chagas' disease.

5'-Nucleotidase↗
PubMed2005

Ala160 and His116 residues are involved in activity and specificity of apyrase, an ATP-hydrolysing enzyme produced by enteroinvasive Escherichia coli.

The virulence plasmid-carried apy (phoN2) gene of Shigella and related enteroinvasive Escherichia coli (EIEC) encodes apyrase, an ATP-diphosphohydrolase belonging to class A of the non-specific acid phosphatases (A-NSAPs). Apyrase and A-NSAPs share three domains of conserved amino acids (domains D1-D3) containing residues forming the putative active site of apyrase. In spite of their similarity, apyrase and A-NSAPs show different substrate specificity, apyrase being able to hydrolyse nucleotide tri- and diphosphates, but not monophosphates, as well as p-nitrophenyl phosphate (pNPP), while A-NSAPs are also active towards monophosphates and pNPP. In this paper, to get further insights into the structure-function relationship of apyrase, a random and site-directed mutagenesis of the apy gene of EIEC strain HN280 was conducted. Results indicate that amino acids located within the D2 and D3 conserved domains (Ser157 and Arg192, respectively) as well as residues located in the N-terminal (Ser97) and C-terminal (Glu233) domains are required for enzyme activity. Surprisingly, Ala160, located near the D2 domain and considered to be important for enzyme specificity, is required for enzyme activity, as its substitution with Thr led to the inactivation of enzyme activity. Furthermore, residue His116 is involved in apyrase specificity, since the H116L apyrase mutant shows substrate specificity resembling that of A-NSAPs.

Adenosine Triphosphate↗
PubMed1999

A Nod factor-binding lectin is a member of a distinct class of apyrases that may be unique to the legumes.

Recent studies from our laboratory have found that a root lectin from the legume Dolichos hifloris is present on the root surface, binds rhizobial Nod factor and has apyrase activity. To assess the broader significance of this lectin/nucleotide phosphohydrolase (Db-LNP), we have cloned a second related cDNA (Db-apyrase-2) from D. hiflorus, as well as related cDNAs from the legumes Lotus japonicus and Medicago sativa, and from Arabidopsis thaliana, a non-legume. The deduced amino acid sequences of these apyrases were aligned with one another and with the sequences of other apyrases from plants, animals, yeast and protozoa. Phylogenetic analysis shows that Db-LNP has closely related orthologs only in other legumes, while Db-apyrase-2 is more closely related to apyrase sequences from non-leguminous plants. We also show that the orthologs of Db-LNP from M. sativa and Pisum sativum have carbohydrate binding activity. The results suggest that legume LNPs may represent a special class of apyrases that arose by gene duplication and subsequent specialization.

Amino Acid Sequence↗
PubMed1996

Apyrase activity and adenosine diphosphate induced platelet aggregation inhibition by the salivary gland proteins of Culicoides variipennis, the North American vector of bluetongue viruses.

Salivary gland homogenates of Culicoides variipennis, the primary vector of bluetongue (BLU) viruses in North America, were analyzed for apyrase activity. Apyrase (ATP diphosphohydrolase, EC 3.6.1.5) is an anti-hemostatic and anti-inflammatory salivary enzyme of most hematophagous arthropods. The enzyme activity was measured by the release of orthophosphate using ATP, ADP, and AMP as substrates with Ca2+ as the divalent cation. ATPase (11.5 +/- 1 mU/pair of glands), ADPase (7.3 +/- 0.7 mU/pair of glands), and insignificant (P < 0.05) AMPase (0.07 mU/pair of glands) activities were detected in female salivary glands. Male salivary glands contained lower amounts of ATPase and ADPase activity (P < 0.05). The ATPase and ADPase activities were greatest at pH 8.5, and were similarly activated by Mg2+. Molecular sieving HPLC of salivary gland homogenates generated a single peak which coincided with ATPase and ADPase, but no AMPase, activity; the protein has an estimated molecular mass of 35,000 Da. ATPase and ADPase activity, and total protein concentration, were reduced (P < 0.05) in the salivary glands of females after taking a blood meal from a sheep. Salivary gland homogenates also inhibited ADP-induced platelet aggregation in vitro. It is concluded that the salivary ATPase and ADPase activities of C. variipennis reside in one enzyme, and that this enzyme is likely an apyrase. The apyrase activity is thought to be responsible for the inhibition of ADP-induced platelet aggregation, as indicated by the apparent discharge of apyrase from salivary glands into the host during blood feeding. This suggests that apyrase is one of the salivary proteins present in C. variipennis acting as antigens in the development of Culicoides hypersensitivity in ruminants and horses. Apyrase may inhibit an inflammatory response at the feeding site through the subsequent degradation of its end-product, AMP, to adenosine, a potent anti-inflammatory substance, by the ecto-5' nucleotidase activity of neutrophils.

Adenosine Diphosphate↗
PubMed2003

Methylotrophic yeast Pichia pastoris as a host for production of ATP-diphosphohydrolase (apyrase) from potato tubers (Solanum tuberosum).

ATP-diphosphohydrolase (apyrase) catalyzes the hydrolysis of phosphoanhydride bonds of nucleoside tri- and di-phosphates in the presence of divalent cations. This enzyme has broad substrate specificity for nucleotides, which makes it an ideal enzyme for different biotechnical applications, such as DNA sequencing and platelet-aggregation inhibition. The only commercially available apyrase is isolated from potato tubers. To avoid batch-to-batch variations in activity and quality, we decided to produce a recombinant enzyme. The methylotrophic yeast Pichia pastoris was chosen as an eukaryotic expression host. The coding sequence of potato apyrase, without the signal peptide, was cloned into the YpDC541 vector to create a fusion with the alpha-mating secretion signal of Saccharomyces cerevisiae. The gene was placed under the control of the methanol-inducible alcohol oxidase promoter. The YpDC541-apyrase construct was integrated into P. pastoris strain SMD1168. Methanol induction resulted in secretion of apyrase to a level of 1mg/L. The biologically active recombinant apyrase was purified by hydrophobic interaction and ion exchange chromatography. According to SDS-PAGE and Western blot analysis, the purified enzyme showed to be hyperglycosylated. By enzymatic removal of N-glycans, a single band corresponding to a molecular mass of 48kDa was detected. The recombinant apyrase was found to function well when it was used in combination with the Pyrosequencing technology.

Alcohol Oxidoreductases↗
PubMed2004

Intravenous apyrase administration reduces arterial thrombosis in a rabbit model of endothelial denudation in vivo.

The role of adenine nucleotides on vascular and platelet functions has long been established. Apyrase (CD39) takes part of a family of ecto-enzymes that hydrolyze adenosine diphosphate and adenosine triphosphate. The participation of apyrase in the thromboregulatory system is under study. An in vivo experimental model of acute arterial thrombosis was used to test the hypothesis that administering a soluble form of potato apyrase could prevent thrombus formation. Twenty-five white New Zealand male rabbits suffered balloon aortic endothelium denudation and, after 15 days, they were submitted to a thrombosis-triggering protocol with a procoagulant (Russel's viper venom) and epinephrine. After the thrombosis-triggering protocol, 12 animals received two soluble apyrase administrations intravenously (with 90 min intervals), while 13 control animals received no apyrase. Three hours after the triggering protocol, the animals were killed and the rate and area of arterial thrombosis were analyzed. The rate of thrombosis in the apyrase group was significantly lower than that of the control group (16.7 versus 69%, respectively; P = 0.015), as was the area of thrombosis (1.7 +/- 4.3 versus 21.7 +/- 37.4 mm2, respectively; P = 0.008). Our results confirm that apyrase participates in homeostasis through a potent anti-thrombotic effect.

Adenine Nucleotides↗