共查询到20条相似文献,搜索用时 15 毫秒
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Mardanyan S Sharoyan S Antonyan A Armenyan A Cristalli G Lupidi G 《Biochimica et biophysica acta》2001,1546(1):185-195
Adenosine deaminase from bovine cerebral hemisphere (white and gray matter) and spleen was treated with N-bromosuccinimide, a reagent known to oxidize selectively tryptophan residues in proteins. Spectrally observable tryptophan modification was accompanied by enzyme inactivation. Tsow graphics revealed that two Trps are essential for the activity of enzyme from both tissues. Enzyme inhibitors and substrate analogues, derivatives of erythro-9-(2-hydroxy-3-nonyl)adenine (EHNA) and adenosine, were able to protect Trp against modification, and this effect correlated in general with the enzyme activity protection. In the presence of adenosine deaza analogues (the noninhibitor tubercidin among them) only two Trps were modified in the fully inactivated enzyme. In the presence of EHNA and its deaza analogues, full inactivation of the enzyme was accompanied by the modification of four Trps. The obtained data confirm the previous hypothesis about the presence on the enzyme of different binding sites for adenosine and EHNA derivatives that are responsible for the different effects on the enzyme conformation elicited by the corresponding derivatives. Moreover, these data allow us to suggest that Trp residues, still unidentified by X-ray analysis, are essential for the functioning of the enzyme. 相似文献
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The enzyme adenosine kinase (AK) plays a central role in regulating the intracellular and interstitial concentration of the
purine nucleoside adenosine (Ado). In view of the beneficial effects of Ado in protecting tissues from ischemia and other
stresses, there is much interest in developing AK inhibitors, which can regulate Ado concentration in a site- and event-specific
manner. The catalytic activity of AK from different sources is dependent upon the presence of activators such as phosphate
(Pi). In this work we describe several new phosphorylated compounds which either activate or inhibit AK. The compounds acetyl
phosphate, carbamoyl phosphate, dihydroxyacetone phosphate and imidodiphosphate were found to stimulate AK activity in a dose-dependent
manner comparable to that seen with Pi. In contrast, a number of phosphonate and bisphosphonate derivatives, which included
clodronate and etidronate, were found to inhibit the activity of purified AK in the presence of Pi. These AK inhibitors (viz. clodronate, etidronate, phosphonoacetic acid, 2-carboxyethylphosphonic acid, N-(phosphonomethyl)-glycine and N-(phosphonomethyl)iminodiacetic acid), at concentrations at which they inhibited AK, were also shown to inhibit the uptake
of 3H-adenosine and its incorporation into macromolecules in cultured mammalian cells, indicating that they were also inhibiting
AK in intact cells. The drug concentrations at which these effects were observed showed limited toxicity to the cultured cells,
indicating that these effects are not caused by cellular toxicity. These results indicate that the enzyme AK provides an additional
cellular target for the clinically widely used bisphosphonates and related compounds, which could possibly be exploited for
a new therapeutic application. Our structure–activity studies on different AK activators and inhibitors also indicate that
all of the AK activating compounds have a higher partial positive charge (δ+) on the central phosphorous atom in comparison to the inhibitors. This information should prove helpful in the design and
synthesis of more potent inhibitors of AK. 相似文献
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Saboury AA Divsalar A Ataie G Amanlou M Moosavi-Movahedi AA Hakimelahi GH 《Acta biochimica Polonica》2003,50(3):849-855
Kinetic and thermodynamic studies were made on the effect of caffeine on the activity of adenosine deaminase in 50 mM sodium phosphate buffer, pH 7.5, using UV spectrophotometry and isothermal titration calorimetry (ITC). An uncompetitive inhibition was observed for caffeine. A graphical fitting method was used for determination of binding constant and enthalpy of inhibitor binding by using isothermal titration microcalorimetry data. The dissociation-binding constant is equal to 350 microM by the microcalorimetry method, which agrees well with the value of 342 microM for the inhibition constant that was obtained from the spectroscopy method. Positive dependence of caffeine binding on temperature indicates a hydrophobic interaction. 相似文献
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J M May 《Journal of cellular physiology》1988,135(2):332-338
The human erythrocyte membrane carriers for hexoses and nucleosides have several structural features in common. In order to assess functional similarities, the effects of adenosine derivatives on hexose transport and cytochalasin B binding sites were studied. Adenosine inhibited zero-trans uptake of 3-O-methylglucose half-maximally at 5 mM, while more hydrophobic adenosine deaminase-resistant derivatives were ten- to 20-fold more potent transport inhibitors. However, degradation of adenosine accounted for very little of this difference in potency. Hexose transport was rapidly inhibited by N6-(L-2-phenylisopropyl)adenosine at 5 degrees C in a dose-dependent fashion (EC50 = 240 microM), to lower the transport Vmax without affecting the Km. A direct interaction with the carrier protein was further indicated by the finding that N6-(L-2-phenylisopropyl)adenosine competitively inhibited [3H]cytochalasin B binding to erythrocytes (Ki = 143 microM) and decreased [3H]cytochalasin B photolabeling of hexose carriers in erythrocyte ghosts. The cross-reactivity of adenosine and several of its derivatives with the hexose carrier suggests further homologies between the carriers for hexoses and nucleosides, possibly related to their ability to transport hydrophilic molecules through the lipid core of the plasma membrane. 相似文献
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J J Centelles R Franco J Bozal 《Comparative biochemistry and physiology. B, Comparative biochemistry》1987,86(1):95-98
The level of adenosine deaminase in various rat tissues has been tested. The enzyme activity of cytosolic fractions decreased in the following order: lung greater than spleen greater than small intestine greater than stomach greater than kidney greater than heart greater than liver greater than skeletal muscle greater than forebrain greater than cerebellum. The enzyme had identical patterns from tissue to tissue with respect to Km, V, and Ki values for ethanol and for dimethyl sulfoxide, with respect to electrophoretic behaviour and to inhibition by antibodies anti-rat brain adenosine deaminase. 相似文献
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Inhibition of adenosine deaminase by a metabolite of the nephrotoxic drug, puromycin aminonucleoside
N Dickie L Norton R F Derr C S Alexander H T Nagasawa 《Proceedings of the Society for Experimental Biology and Medicine. Society for Experimental Biology and Medicine (New York, N.Y.)》1966,123(2):421-423
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W P Schrader C A West N L Strominger 《The journal of histochemistry and cytochemistry》1987,35(4):443-451
Adenosine deaminase and adenosine deaminase complexing protein have been localized in rabbit brain. Brains fixed in paraformaldehyde or in Clarke's solution were blocked coronally. Blocks from brains fixed in paraformaldehyde were either frozen in liquid nitrogen or embedded in paraffin. Tissue fixed in Clarke's solution was embedded in paraffin. Sections from each block were stained by the peroxidase-antiperoxidase method for adenosine deaminase or complexing protein using affinity-purified goat antibodies. Adenosine deaminase and complexing protein did not co-localize. Adenosine deaminase was detected in oligodendroglia and in endothelial cells lining blood vessels, whereas complexing protein was concentrated in neurons. The subcellular location and appearance of the peroxidase reaction product associated with individual cells was also quite distinctive. The cell bodies of adenosine deaminase-positive oligodendroglia were filled with intense deposits of peroxidase reaction product. In contrast to oligodendroglia, the reaction product associated with most neurons stained for complexing protein was concentrated in granular-appearing cytoplasmic deposits. In some instances, these deposits were clustered about the nuclear membrane. Staining of neurons in the granular layer of cerebellum was an exception. Granule cells were lightly outlined by peroxidase reaction product. Cerebellar islands, also referred to as glomeruli, were stained an intense uniform brown. These results raise the possibility that oligodendroglia and blood vessel endothelia, through the action of adenosine deaminase, might play a role in controlling the concentration of extracellular adenosine in brain. They do not, however, support the suggestion that complexing protein aids in adenosine metabolism by positioning adenosine deaminase on the plasma membrane. 相似文献
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Adenosine deaminase (adenosine aminohydrolase; E.C. 3.5.4.4) has been purified 4686-fold from egg yolk. The procedure developed was used to isolate the enzyme from eight chicken eggs. An easily prepared affinity column employing purine riboside was used as the final step in the purification. The method developed permits the rapid isolation and a high recovery of the protein. The specific activity of the enzyme preparation obtained is 81.4 mU/mg. 相似文献
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Adenosine deaminase (ADA) was isolated from small intestine of mice and purified to utmost homogeneity. SDS-PAGE of purified ADA gave a molecular weight of 41 kDa. Western blot analyses gave a single reactive band at 41 kDa and the other band was an associated ADA binding protein. The purified enzyme was more stable in the alkaline pH. The optimum pH and the pI values were about 7.0 and 4.96, respectively. Km values of the small intestinal ADA for adenosine and 2-deoxyadenosine were 23 and 16M, respectively. Purine riboside was a competitive inhibitor with Ki of 5 M, whereas 2-3-o-isopropylidene adenosine acted as an uncompetitive inhibitor (Ki 66 M). Activity of ADA was inhibited by the presence of theophylline (-40%), caffeine (-30%), and L-cysteine (-50%). Significantly, Hg2+ (100 M) inhibited 98% of the initial ADA activity. In addition, various purine analogs such as inosine, purine, -adenosine and adenine showed variable inhibitions on the activity of ADA. Relative ADA activity towards 3-deoxyadenosine and 6-chloropurine riboside was lower by 30% and 40%, respectively. However, the activity towards 2-o-methyl adenosine was higher (30%) compared to the activity obtained using adenosine. 相似文献
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Inhibition of adenosine deaminase activity leads to decreased cellular immunity. The effect of deoxycoformycin (DCF), a potent inhibitor of adenosine deaminase, on the ability of mouse spleen cells to generate antibody responses in vitro has been examined. With either continuous exposure to or pretreatment of the cells with deoxycoformycin, there was a decrease in cell survival and an increase in antibody-producing cells in the surviving cell population. To identify the cell population most susceptible to the inhibitor, the spleen was separated into B-cell, and T-cell, and macrophage components and each population was pretreated with deoxycoformycin before combination with its complementary treated or untreated population. Deoxycoformycin pretreatment had no effect on macrophages or B cells; however, pretreatment of the T cells resulted in increased antibody responses. When T cells and B cells were both pretreated and combined, there was a synergistic increase in the antibody response. In addition, supernatants from cultures in which both B cells and T cells had been pretreated with DCF were capable of enhancing antibody responses in cultures containing DCF-treated T cells. Though adenosine was increased in the stimulatory culture supernatants, adenosine alone did not enhance antibody responses in either untreated or DCF-treated cultures. 相似文献
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