Sequencing, functional expression and characterization of rat NTPDase6, a nucleoside diphosphatase and novel member of the ecto-nucleoside triphosphate diphosphohydrolase family

Braun, Norbert; Fengler, Sabine; Ebeling, Christian; Servos, Jörg; Zimmermann, Herbert

doi:10.1042/0264-6021:3510639

Cited by 44 publications

(54 citation statements)

References 34 publications

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“…The two isoforms of human NTPDase4 hydrolyze nucleoside tri-and diphosphates, but ATP and ADP only to a minor extent [63,76]. Both NTPDase5 and NTPDase6 reveal a high preference for nucleoside diphosphates whereby ADP is a poor substrate [39,[77][78][79]. NTPDase7 preferentially hydrolyses UTP, GTP, and CTP, but ATP and nucleoside diphosphates only to a minor extent [64].…”

Section: General Properties and Functional Rolementioning

confidence: 99%

Cellular function and molecular structure of ecto-nucleotidases

Zimmermann

Zebisch

Sträter

2012

Purinergic Signalling

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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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Section: General Properties and Functional Rolementioning

confidence: 99%

Cellular function and molecular structure of ecto-nucleotidases

Zimmermann

Zebisch

Sträter

2012

Purinergic Signalling

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922

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“…In contrast, analysis of the amino acid sequence of SmATPDase2 shows that it more closely resembles intracellular and soluble enzymes of the NTPDase 5 and 6 families [18]. Proteins in these families have been reported to support glycosylation reactions in the Golgi apparatus or the endoplasmic reticulum [30][31][32]. It is possible that SmATPDase2 likewise participates in Golgi and/ or ER glycosylation reactions inside schistosomes.…”

Section: Discussionmentioning

confidence: 97%

Schistosome apyrase SmATPDase1, but not SmATPDase2, hydrolyses exogenous ATP and ADP

Da’dara¹,

Bhardwaj²,

Skelly³

2014

Purinergic Signalling

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Schistosomes are parasitic worms that can live in the bloodstream of their vertebrate hosts for many years. It has been proposed that the worms impinge on host purinergic signalling by degrading proinflammatory molecules like ATP as well as prothrombotic mediators like ADP. This capability may help explain the apparent refractoriness of the worms to both immune elimination and thrombus formation. Three distinct ectoenzymes, expressed at the host-exposed surface of the worm's tegument, are proposed to be involved in the catabolism of ATP and ADP. These are alkaline phosphatase (SmAP), phosphodiesterase (SmNPP-5), and ATP diphosphohydrolase (SmATPDase1). It has recently been shown that only one of these enzymes-SmATPDase1-actually degrades exogenous ATP and ADP. However, a second ATP diphosphohydrolase homolog (SmATPDase2) is located in the tegument and has been reported to be released by the worms. It is possible that this enzyme too participates in the cleavage of exogenous nucleotide tri-and di-phosphates. To test this hypothesis, we employed RNA interference (RNAi) to suppress the expression of the schistosome SmATPDase1 and SmATPDase2 genes. We find that only SmATPDase1-suppressed parasites are significantly impaired in their ability to degrade exogenously added ATP or ADP. Suppression of SmATPDase2 does not appreciably affect the worms' ability to catabolize ATP or ADP. Furthermore, we detect no evidence for the secretion or release of an ATP-hydrolyzing activity by cultured parasites. The results confirm the role of tegumental SmATPDase1, but not SmADTPDase2, in the degradation of the exogenous proinflammatory and prothrombotic nucleotides ATP and ADP by live intravascular stages of the parasite.

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“…However, several family members exhibit a topology commonly observed among channels, transporters, and receptors but unusual for ectoenzymes: a large extracellular domain flanked by two transmembrane helices (9)(10)(11)20,21). Why the active site would be held down by transmembrane domains on both sides rather than by a single protein or lipid link was originally a mystery, but in recent years it has become clear that the two transmembrane helices are intricately linked to active site function rather than simply serving as anchors (22).…”

Section: Nih-pa Author Manuscriptmentioning

confidence: 99%

Bilayer Mechanical Properties Regulate the Transmembrane Helix Mobility and Enzymatic State of CD39

Grinthal

Guidotti

2006

Biochemistry

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CD39 can exist in at least two distinct functional states depending on the presence and intact membrane integration of its two transmembrane helices. In native membranes, the transmembrane helices undergo dynamic rotational motions that are required for enzymatic activity and are regulated by substrate binding. In the present study we show that bilayer mechanical properties regulate conversion between the two enzymatic functional states by modulating transmembrane helix dynamics. Alteration of membrane properties by insertion of cone shaped or inverse cone shaped amphiphiles or by cholesterol removal switches CD39 to the same enzymatic state as does removing or solubilizing the transmembrane domains. The same membrane alterations increase the propensity of both transmembrane helices to rotate within the packed structure, resulting in a structure with greater mobility but not an altered primary conformation. Membrane alteration also abolishes the ability of substrate to stabilize the helices in their primary conformation, indicating a loss of coupling between substrate binding and transmembrane helix dynamics. Removal of either transmembrane helix mimics the effect of membrane alteration on the mobility and substrate sensitivity of the remaining helix, suggesting that the ends of the extracellular domain have intrinsic flexibility. We suggest that a mechanical bilayer property, potentially elasticity, regulates CD39 by altering the balance between stability and flexibility of its transmembrane helices and, in turn, of its active site. KeywordsCD39; NTPDase; transmembrane helices; bilayer; mechanical Of the various enzymes that process nucleotides at the cell surface and in the lumen of intracellular organelles, the ectonucleoside triphosphate diphosphohydrolases (eNTPDases) have emerged as the major family responsible and specific for breaking the terminal phosphoanhydride bonds of tri-and dinucleotides (1,2). As such, they have been shown or hypothesized to modulate many of the signaling and biosynthetic processes in which extracytoplasmic nucleotides play a role, including vascular homeostasis, cell size maintenance, neuronal signaling, immune function, and protein and lipid modification (3)(4)(5)(6)(7)(8). Consistent with the variety of tasks they perform on and in the cell, different family members exhibit different localizations and specificities: some reside on the plasma membrane and others in the Golgi, lysosomes, or endoplasmic reticulum, while each has a characteristic hierarchy of preferences for di-vs trinucleotides as well as for different bases (1,9-11). The defining structural feature shared by all family members is a set of five short sequences called apyrase conserved regions (ACRs) (12,13), two of which are thought to constitute phosphate binding loops based on homology to the nucleotide binding domain of the actin/hsp70/ † This work was supported by Grant HL08893 from the National Institutes of Health.*To whom correspondence should be addressed at 7 Divinity Ave., Cambridge, MA 02138. P...

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Sequencing, functional expression and characterization of rat NTPDase6, a nucleoside diphosphatase and novel member of the ecto-nucleoside triphosphate diphosphohydrolase family

Cited by 44 publications

References 34 publications

Cellular function and molecular structure of ecto-nucleotidases

Cellular function and molecular structure of ecto-nucleotidases

Schistosome apyrase SmATPDase1, but not SmATPDase2, hydrolyses exogenous ATP and ADP

Bilayer Mechanical Properties Regulate the Transmembrane Helix Mobility and Enzymatic State of CD39

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