Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

---

Query: EC:3.6.1.25 (triphosphatase)
1,529 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have investigated the stereoselectivity of ectonucleotidases (nucleoside triphosphatase, EC 3.6.1.15; nucleoside diphosphatase, EC 3.6.1.6; 5'-nucleotidase, EC 3.1.3.5) on pig aortic endothelial cells using two classes of nucleotide analogue. In experiments with nucleotide enantiomers in which the natural D-ribofuranosyl moiety is replaced by an L-ribofuranosyl moiety, the rate of catabolism of 100 microM-L-ATP was one-fifth that of D-ATP, the rate of catabolism of 100 microM-L-ADP was one-fifteenth that of D-ADP and there was no detectable catabolism of 100 microM-L-AMP. Each of the L-enantiomers inhibited, apparently competitively, the catabolism of the corresponding D-enantiomer; Ki values were approx. 0.6 mM, 1.0 mM and 3.9 mM for L-ATP, L-ADP and L-AMP respectively. Experiments with adenosine 5'-[beta, gamma-imido]triphosphate and with D- and L-enantiomers of adenosine 5'-[beta, gamma-methylene]triphosphate revealed modest ectopyrophosphatase activity, undetectable in experiments with natural nucleotides, which was also stereoselective. Use of phosphorothioate nucleotide analogues demonstrated that ATP catabolism was virtually stereospecific with respect to the geometry of the thiol group substituted on the beta-phosphate: the Rp isomer was degraded, whereas there was little or no breakdown of the Sp isomer. ADP catabolism was also stereospecific with respect to the geometry of the thiol group substituted on the alpha-phosphate: the Sp isomer but not the Rp isomer was degraded. The geometry of thiol-group substitution on the alpha-phosphate had no effect on ATP catabolism to ADP. There was no detectable catabolism of analogues with thiol-group substitution on the terminal phosphate. Each of the phosphorothioate analogues that was catabolized broke down at a rate similar to that of the natural nucleotide from which it was derived. These results demonstrate that the ectonucleotidases on pig aortic endothelial cells exhibit a high degree of stereoselectivity, characteristic for each enzyme, both with respect to the ribofuranosyl moiety and to the phosphate side chain.
...
PMID:Stereoselectivity of ectonucleotidases on vascular endothelial cells. 631 68

The kinetics of nucleoside-triphosphate-dependent proton translocation by a vacuolar-type adenosine-triphosphatase have been studied, using the enzyme from bovine chromaffin-granule membranes, purified and reconstituted into proteoliposomes. The reaction was followed by recording the quenching of the fluorescence of the permeant weak base 9-amino-6-chloro-2-methoxyacridine; fluorescence data were collected and stored in digital form, and the initial reaction rates estimated by linear regression. In the absence of nucleoside diphosphate, the dependence of initial rates of proton translocation on substrate concentration were fitted well by the Michaelis-Menten equation, as were the kinetics of ATP hydrolysis. ADP and other nucleoside diphosphates were potent inhibitors of the ATPase, effecting a reduction in the maximum velocity of the reaction, and producing sigmoid substrate-saturation curves which could be fitted by the empirical Hill equation, the Hill coefficient approaching 2 at high inhibitor concentrations. Data sets containing initial-rate estimates were collected over a wide range of independently varied concentrations of substrate and inhibitor and were modeled, using rate equations derived from several different models based on the concerted-transition model of allosteric inhibition proposed by Monod, Wyman and Changeux. These equations were fitted to the data by weighted non-linear regression, using an iterative computer program to obtain the best estimates of kinetic parameters. One model consistently fitted all of the data sets better than all the others, and this model was based on the following assumptions: that the ATPase exists in two conformational states, R and T; that only the R state is catalytically active; that each state contains three kinetically equivalent catalytic sites, and one regulatory site; that nucleoside triphosphates bind only to the catalytic sites, and that nucleoside diphosphates bind both to the catalytic sites and to the regulatory site. The optimized values of the kinetic parameters indicate that in the absence of nucleoside diphosphate, the enzyme is almost completely in the R state; that nucleoside triphosphates bind more tightly to the R than to the T state; that binding of nucleoside diphosphates to the regulatory site is very tight, but occurs only in the T state; and that competitive binding of nucleoside diphosphates at the catalytic sites is stronger in the T state than in the R state. Experiments conducted with varying total magnesium concentrations indicated that the magnesium complexes of nucleoside diphosphates are much stronger inhibitors than the free nucleotides, and that free nucleoside triphosphates are weakly inhibitory, probably competing with the magnesium complexes for binding at the catalytic sites.(ABSTRACT TRUNCATED AT 400 WORDS)
...
PMID:Allosteric regulation of proton translocation by a vacuolar adenosinetriphosphatase. 755 11

Sedimentation and high performance liquid chromatography studies show that the functional DNA replication helicase of bacteriophage T4 (gp41) exists primarily as a dimer at physiological protein concentrations, assembling from gp41 monomers with an association constant of approximately 10(6) M-1. Cryoelectron microscopy, analytical ultracentrifugation, and protein-protein cross-linking studies demonstrate that the binding of ATP or GTP drives the assembly of these dimers into monodisperse hexameric complexes, which redissociate following depletion of the purine nucleotide triphosphatase (PuTP) substrates by the DNA-stimulated PuTPase activity of the helicase. The hexameric state of gp41 can be stabilized for detailed study by the addition of the nonhydrolyzable PuTP analogs ATP gamma S and GTP gamma S and is not significantly affected by the presence of ADP, GDP, or single-stranded or forked DNA template constructs, although some structural details of the hexameric complex may be altered by DNA binding. Our results also indicate that the active gp41 helicase exists as a hexagonal trimer of asymmetric dimers, and that the hexamer is probably characterized by D3 symmetry. The assembly pathway of the gp41 helicase has been analyzed, and its structure and properties compared with those of other helicases involved in a variety of cellular processes. Functional implications of such structural organization are also considered.
...
PMID:The phage T4-coded DNA replication helicase (gp41) forms a hexamer upon activation by nucleoside triphosphate. 770 92

To directly assess the possible role of ADP in muscle fatigue, we have studied the effect of physiological MgADP levels on maximum Ca(2+)-activated isometric force and unloaded shortening velocity (Vus) of single skinned fiber segments from rabbit fast-twitch (psoas) and slow-twitch (soleus) muscles. MgADP concentration was changed in a controlled and well-buffered manner by varying creatine (Cr) in solutions, which also contained MgATP, phosphocreatine (PCr), and creatine kinase (CK). To quantify ADP as a function of Cr added, we determined the apparent equilibrium constant (K') of CK for the conditions of our experiments (pH 7.1, 3 mM Mg2+, 12 degrees C): K' = (sigma [Cr]. sigma [ATP])/(sigma [PCr]. sigma [ADP]) = 260 +/- 3 (SE). In this manner, ADP was altered essentially as occurs during stimulation in vivo but without the concomitant changes in pH and P(i), which affect force and Vus. As ADP (and Cr) was increased, force and Vus decreased in both fiber types; at the highest ADP level used, 200 microM, normalized force was 96.6 +/- 1.7% for psoas (n = 6) and 93.7 +/- 2.8% for soleus (n = 6), and Vus was 80.4 +/- 2.4% for psoas and 91.3 +/- 7.7% for soleus. Diffusion-reaction calculations indicated that radial gradients of metabolite concentrations within fibers could not explain the small effects of ADP on fiber mechanics, and experiments verified that metabolite levels were well buffered within fibers by the CK reaction. Exogenous CK was added to bathing solutions at 290 U/ml, threefold above that necessary to maintain Vus independent of CK concentration; in the absence of PCr and exogenous CK, at least a fourfold increased MgATP was necessary to maintain Vus at the control level. Adenylate kinase activity was not detectable; thus myofibrillar adenosine-triphosphatase and exogenous CK activities were the major determinants of nucleotide levels within activated cells. Cr alone (in absence of PCr and exogenous CK) also decreased force and Vus, presumably by a nonspecific mechanism. Over the physiological range, altered ADP had little or no effect on force or Vus in well-buffered conditions. It is therefore likely that other factors decrease force and Vus during muscular fatigue.
...
PMID:Effect of physiological ADP concentrations on contraction of single skinned fibers from rabbit fast and slow muscles. 786 87

1. Parotid plasma membrane nonpump low-affinity Ca(2+)-ATPase, which possesses high-affinity (Ca2+ + Mg2+)-ATPase activity, was characterized. 2. Purified Ca(2+)-ATPase hydrolyzed the nucleoside triphosphates, GTP, ITP, CTP, UTP, TTP (67-93% of ATP) and nucleoside diphosphates, ADP, GDP, IDP, CDP, TDP (12-40% of ATP) but not AMP and p-NPP. 3. The maximum activities of Ca(2+)- and (Ca2+ + Mg2+)-ATPases were obtained in the presence of 1 mM and 0.13 microM Ca2+, respectively. 4. The Km values for Ca2+ in Ca(2+)- and (Ca2+ + Mg2+)-ATPases were 0.2 mM and 22 nM, respectively. 5. The activities of both Ca(2+)- and (Ca2+ + Mg2+)-ATPases were found in the right-side-out-vesicles obtained from the plasma membrane-rich fraction. 6. These features suggest that Ca(2+)-ATPase is an ecto-Ca(2+)-dependent nucleoside triphosphatase.
...
PMID:The possibility that Ca(2+)-ATPase from the plasma membrane-rich fraction of bovine parotid gland is ecto-Ca(2+)-dependent nucleoside triphosphatase. 806 15

The relationship between ecto-ATPase activity and the vasoactive effect of ATP is unclear. Previously we have characterized the ectonucleoside triphosphatase activity of isolated rat mesenteric small arteries and now characterize the effect of nucleotides on the tone of these arteries. In resting arteries, ATP caused concentration-dependent contractions that were transient and could not be reproduced within 2 h. Transient contractions in response to ATP also were elicited in arteries precontracted with norepinephrine, but the potency of ATP was increased and responses to repeated stimulations could be obtained. Contractions were followed by relaxation. The response to ATP was unaffected by 100 microM theophylline, 1 microM propranolol or removal of the endothelium. Transient contractions followed by relaxation were caused also by ADP, 2-methyl-thio-ATP (2meSATP) and alpha, beta-methylene-ATP (alpha, beta-meATP). UTP caused sustained contractions, whereas GTP and ITP had little effect. The rank order of potency (alpha, beta-,mATP > ATP > ADP) suggested that P2x purinoceptors were responsible for the contractions, whereas the rank order of potency for the relaxation (alpha, beta-meATP > or = ATP > 2meSATP) was not consistent with the relaxation being mediated by P2Y purinoceptors as defined originally. Desensitization of the contractile response to ATP by alpha, beta-meATP was variable. In contrast, inhibition of the response to ATP was obtained consistently and dose-dependently with GTP.
...
PMID:Effects of ATP and related nucleotides on the tone of isolated rat mesenteric resistance arteries. 845 Apr 60

We investigated in vitro motility of F-actin on heavy meromyosin (HMM) and nucleotide triphosphatase activity of acto-HMM by using ATP analogues of various nucleotide triphosphates (NTPs) and enzymatically cleaved actins. The sliding velocity did not correlate with the actin activated HMM-NTPase activity, but correlated strongly with the reciprocal of NTPase activity of HMM itself, i.e., the cycle time of HMM NTPase. This indicated that with ATP the complex of myosin with the product, M.ADP.Pi, at the long lived intermediate state of the rate limiting step would play a key role for efficient mechanochemical energy transduction during actin-myosin interaction.
...
PMID:The mechanism for mechanochemical energy transduction in actin-myosin interaction revealed by in vitro motility assay with ATP analogues. 863 37

We have described a thyroid hormone receptor in synaptosomes of the chick embryo brain. To understand how the hormones exert their actions at this level, we performed a series of studies to demonstrate that this receptor could be linked to G proteins. Guanosine 5'-[gamma-thio]triphosphate (GTP gamma S)(100 muM) lowered the binding capacity of the receptor high affinity site from 8.9 +/- 1.3 to 3.4 +/- 1.3 ng T3/mg protein, a finding consistent with the coupling of receptor to G proteins. Furthermore, ADP ribosylation with pertussis toxin showed that thyroid hormones induced a dose-dependent increase in the inactive alpha 0-subunit of the G0 protein. This effect was detected at 10 pM, with a maximal increase (mean +/- SEM, 50 +/- 3.6%) at 100 nM, and T4 was as effective as T3. Both hormones also decreased the intrinsic guanine triphosphatase activity of G proteins by lowering the binding of GTP to the alpha-subunit and their rate of hydrolysis. This inhibition was greater with T4 (25 +/- 5%) than with T3 (14 +/- 2%), suggesting that the former could be the more active hormone at the synaptosomal level. The effect on guanine triphosphatase activity confirms that the synaptosomal thyroid hormone receptor is coupled to a G(zero) protein. These results demonstrate that thyroid hormones increase or favor the ADP ribosylation of G alpha(zero) by pertussis toxin. Thus, they enhance the alpha(zero)-GDP form of the G(zero) protein, namely its inactive conformation. By decreasing the activity of this protein, these hormones may modulate the formation of second messengers in synaptosomes and intervene in the regulation of neuronal proliferation and differentiation induced by several factors. Therefore, thyroid hormones may exert their action on brain maturation at least in part by modulating G alpha(zero) through their synaptosomal receptor.
...
PMID:Effect of thyroid hormones on G proteins in synaptosomes of chick embryo. 864 Dec 9

Nucleotides such as ATP, ADP, UTP or the diadenosine polyphosphates and possibly even NAD+ are extracellular signaling substances in the brain and in other tissues. Enzymes located on the cell surface catalyze the hydrolysis of these compounds and thus limit their spatio-temporal activity. As a final hydrolysis product they generate the nucleoside and phosphate. The paper discusses the biochemical properties, cellular localization and functional properties of surface-located enzymes that hydrolyse nucleotides released from nervous tissue. This is preceded by a brief discussion of nucleotide receptors, cellular storage and mechanisms of nucleotide release. In nervous tissue nucleoside 5'-triphosphates are hydrolysed by ecto-ATP-diphosphohydrolase and possibly in addition also by ecto-nucleoside triphosphatase and ecto-nucleoside diphosphatase. The molecular identity of the ATP-diphosphohydrolase has now been revealed. The hydrolysis of nucleoside 5'-monophosphates is catalysed by 5'-nucleotidase whose biochemical properties and molecular structure have been studied in detail. Little is known about the molecular properties of the diadenosine polyphosphatases. Surface located enzymes for the extracellular hydrolysis of NAD+ and also ecto-protein kinases are discussed briefly. The cellular localization of the ecto-nucleotidases is only partly defined. Whereas in adult mammalian brain activity for hydrolysis of ATP and ADP may be associated with nerve cells or glial cells 5'-nucleotidase appears to have a preferential glial allocation in the adult mammal. The extracellular hydrolysis of the nucleotides is of functional importance not only during synaptic transmission where it functions in signal elimination. It plays a crucial role also for the survival and differentiation of neural cells in vitro and presumably during neuronal development in vivo.
...
PMID:Biochemistry, localization and functional roles of ecto-nucleotidases in the nervous system. 891 94

Occupancy of oxytocin receptor (OTR) binding sites in pregnant rat myometrial membranes with iodinated oxytocin antagonist (OTA), followed by detergent solubilization and size selection, showed that radioactivity eluted in two distinct peaks: one corresponding in size to the isolated receptor (approximately 60 kDa) and the other ranging from 240 to 320 kDa. The unliganded 240- to 320-kDa fraction contained OTRs coupled to G proteins, as the addition of oxytocin (OT) increased guanosine 35S-labeled 5'-O-(3-thiotriphosphate) binding up to twofold in a dose-dependent manner. The effects of OT were blocked by coincubation with OTA. G protein alpha-subunits associated with OTRs in the 240- to 320-kDa peak were identified by immunoadsorption. Significant amounts of both G alpha q/11 and G alpha i3 were associated with the OTR; a lesser amount of G alpha s was complexed. Using the same approach but with antibodies to effector enzymes, we observed that phospholipase C beta 1 (PLC beta 1) and PLA2 were also associated with the OTR. The results corroborate the well-established interaction of OTR with Gq and further show that Gi coupling might be an important component of OTR signal transduction. To further investigate the interaction of Gi with the OTR, we showed that OT stimulation of guanosine 5'-triphosphatase activity in intact myometrial membranes was inhibited by pertussis toxin. Pertussis toxin-stimulated ADP ribosylation of G alpha i in myometrial membranes was also decreased by OT treatment. These findings with pertussis toxin strongly indicate that OTR is coupled to Gi in rat myometrial membranes. The 60-kDa OTR peak (noncoupled receptor) was demonstrable in the myometrium only before the end of gestation and after parturition and accounted for about one-half the 125I-OTA binding activity. At term, there was about a fivefold increase in binding and almost a complete shift to the 240- to 320-kDa-size complex. Thus the established increased sensitivity of the myometrium to OT at term could be the result of both upregulation of OTRs and an increase in the fraction of receptors coupled to signal transduction components, one of which is Gi.
...
PMID:Coupling of oxytocin receptor to G proteins in rat myometrium during labor: Gi receptor interaction. 917 88


<< Previous 1 2 3 4 5 6 Next >>

---