Purification and Properties of Human Placental ATP Diphosphohydrolase

Christoforidis, Savvas; Papamarcaki, Thomais; Galaris, Dimitrios; Kellner, Roland; Tsolas, Orestes

doi:10.1111/j.1432-1033.1995.066_c.x

Cited by 84 publications

(58 citation statements)

References 44 publications

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“…Measurements of their surface activities over a wide range of nucleotide concentrations suggest that NTPDase1 predominates (40%) under physiological conditions (Ͻ1 M ATP), whereas NTPDase3 predominates (70%) in the presence of excess nucleotides, as generated by cell lysis. These findings are consistent with their biochemical properties, as NTPDase1 was characterized as a low-capacity, high-affinity enzyme (11) and NTPDase3 as a high-capacity, low-affinity enzyme (72). Their contributions to the regulation of P2Y 2 R agonists are expected to vary along the airways and over time with the nucleotide concentrations.…”

Section: Discussionsupporting

confidence: 72%

Cystic fibrosis remodels the regulation of purinergic signaling by NTPDase1 (CD39) and NTPDase3

Fausther

Pelletier

Ribeiro

et al. 2010

American Journal of Physiology-Lung Cellular and Molecular Phys

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Airway defenses are regulated by a complex purinergic signaling network located on the epithelial surfaces, where ATP stimulates the clearance of mucin and pathogens. The present study shows that the obstructive disease cystic fibrosis (CF) affects the activity, expression, and tissue distribution of two ectonucleotidases found critical for the regulation of ATP on airway surfaces: NTPDase1 and NTPDase3. Functional polarities and mRNA expression levels were determined on primary cultures of human bronchial epithelial (HBE) cells from healthy donors and CF patients. The in vitro model of the disease was completed by exposing CF HBE cultures for 4 days to supernatant of the mucopurulent material (SMM) collected from the airways of CF patients. We report that NTPDase1 and NTPDase3 are coexpressed on HBE cultures, where they regulate physiological and excess nucleotide concentrations, respectively. In aseptic conditions, CF epithelia exhibit >50% lower NTPDase1 activity, protein, and mRNA levels than normal epithelia, whereas these parameters are threefold higher for NTPDase3. Exposure to SMM induced opposite polarity shifts of the two NTPDases on both normal and CF epithelia, apical NTPDase1 being mobilized to basolateral surfaces and bilateral NTPDase3 to the apical surface. Their immunolocalization in human tissue revealed that NTPDase1 is expressed in epithelial, inflammatory, and endothelial cells, whereas NTPDase3 is restricted to epithelial cells. Furthermore, the SMM-exposed CF HBE cultures reproduced the impact of the disease on their in vivo distribution. This study provides evidence that an extensive remodeling of the enzymatic network regulating clearance occurs in the airways of CF patients.

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Section: Discussionsupporting

confidence: 72%

Cystic fibrosis remodels the regulation of purinergic signaling by NTPDase1 (CD39) and NTPDase3

Fausther

Pelletier

Ribeiro

et al. 2010

American Journal of Physiology-Lung Cellular and Molecular Phys

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“…Polyclonal anti-CD39 antibodies were produced against full-length human placental CD39. The enzyme, 600 g, was isolated from human term placenta (27) and further purified by preparative SDS-PAGE to avoid any contaminants. The final preparation was used for the immunization of two rabbits.…”

Section: Methodsmentioning

confidence: 99%

Cholesterol-dependent Lipid Assemblies Regulate the Activity of the Ecto-nucleotidase CD39

Papanikolaou¹,

Papafotika²,

Murphy³

et al. 2005

Journal of Biological Chemistry

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CD39 (ecto-nucleoside triphosphate diphosphohydrolase-1; E-NTPDase1) is a plasma membrane ecto-enzyme that regulates purinergic receptor signaling by controlling the levels of extracellular nucleotides. In blood vessels this enzyme exhibits a thromboregulatory role through the control of platelet aggregation. CD39 is localized in caveolae, which are plasma membrane invaginations with distinct lipid composition, similar to dynamic lipid microdomains, called rafts. Cholesterol is enriched together with sphingolipids in both rafts and caveolae, as well as in other specialized domains of the membrane, and plays a key role in their function. Here, we examine the potential role of cholesterol-enriched domains in CD39 function. Using polarized MadinDarby canine kidney (MDCK) cells and caveolin-1 genedisrupted mice, we show that caveolae are not essential either for the enzymatic activity of CD39 or for its targeting to plasma membrane. On the other hand, flotation experiments using detergent-free or detergentbased approaches indicate that CD39 associates, at least in part, with distinct lipid assemblies. In the apical membrane of MDCK cells, which lacks caveolae, CD39 is localized in microvilli, which are also cholesterol and raft-dependent membrane domains. Interfering with cholesterol levels using drugs that either deplete or sequester membrane cholesterol results in a strong inhibition of the enzymatic and anti-platelet activity of CD39. The effects of cholesterol depletion are completely reversed by replenishment of membranes with pure cholesterol, but not by cholestenone. These data suggest a functional link between the localization of CD39 in cholesterol-rich domains of the membrane and its role in thromboregulation.

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“…Considerable homology to an intracellular guanosine diphosphatase from yeast was recognized. In parallel, NTPDase1 was purified to homogeneity from human placenta [87]. Partial sequence identification Idealized patterns of nucleotide hydrolysis and product formation by select members of the E-NTPDase, E-NPP, and AP families.…”

Section: General Molecular Propertiesmentioning

confidence: 99%

Cellular function and molecular structure of ecto-nucleotidases

Zimmermann

Zebisch

Sträter

2012

Purinergic Signalling

890

922

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Ecto-nucleotidases play a pivotal role in purinergic signal transmission. They hydrolyze extracellular nucleotides and thus can control their availability at purinergic P2 receptors. They generate extracellular nucleosides for cellular reuptake and salvage via nucleoside transporters of the plasma membrane. The extracellular adenosine formed acts as an agonist of purinergic P1 receptors. They also can produce and hydrolyze extracellular inorganic pyrophosphate that is of major relevance in the control of bone mineralization. This review discusses and compares four major groups of ectonucleotidases: the ecto-nucleoside triphosphate diphosphohydrolases, ecto-5′-nucleotidase, ecto-nucleotide pyrophosphatase/phosphodiesterases, and alkaline phosphatases. Only recently and based on crystal structures, detailed information regarding the spatial structures and catalytic mechanisms has become available for members of these four ecto-nucleotidase families. This permits detailed predictions of their catalytic mechanisms and a comparison between the individual enzyme groups. The review focuses on the principal biochemical, cell biological, catalytic, and structural properties of the enzymes and provides brief reference to tissue distribution, and physiological and pathophysiological functions.

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Purification and Properties of Human Placental ATP Diphosphohydrolase

Cited by 84 publications

References 44 publications

Cystic fibrosis remodels the regulation of purinergic signaling by NTPDase1 (CD39) and NTPDase3

Cystic fibrosis remodels the regulation of purinergic signaling by NTPDase1 (CD39) and NTPDase3

Cholesterol-dependent Lipid Assemblies Regulate the Activity of the Ecto-nucleotidase CD39

Cellular function and molecular structure of ecto-nucleotidases

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