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

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Query: EC:3.5.4.4 (adenosine deaminase)
5,136 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

1. Glutamate inhibits the electrically evoked release of noradrenaline in rabbit brain cortex slices; the inhibition is mediated by adenyl compounds, presumably adenosine. The aim of the present study was to identify the receptors involved in this indirect inhibitory effect of glutamate. Slices of the occipitoparietal cortex were preincubated with [3H]-noradrenaline and then superfused and stimulated by trains of 6 pulses, 100 Hz. 2. The ionotropic glutamate receptor agonists alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AM-PA; 10-100 microM), kainate (10-100 microM) and N-methyl-D-aspartate (NMDA; 30-300 microM) but not the metabotropic glutamate receptor agonist, 1-amino-1,3-cyclopentanedicarboxylate (ACPD; 10-100 microM) reduced the electrically evoked overflow of tritium. 3. The effects of AMPA, kainate and NMDA were attenuated or abolished by the adenosine A1-receptor antagonist, 8-cyclopentyl-1,3-dipropylxanthine (DPCPX) as well as by adenosine A1-receptor antagonist, 8-cyclopentyl-1,3-dipropylxanthine (DPCPX) as well as by adenosine deaminase but not by the alpha 2-adrenoceptor antagonist yohimbine, the gamma-aminobutyric acid (GABA) receptor antagonists, bicuculline and 2-hydroxysaclofen and the NO synthase inhibitor NG-nitro-L-arginine methyl ester (L-NAME). 4. The NMDA receptor antagonist, 2-amino-5-phosphonopentanoate (AP5) blocked the inhibitory effect of NMDA but not that of AMPA and kainate. The non-NMDA-receptor antagonist, 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) blocked the effect of AMPA but not of kainate and NMDA. 5. In addition to decreasing the electrically evoked overflow of tritium, AMPA, kainate and NMDA but not ACPD caused a steep but transient rise of basal tritium efflux. This immediate releasing effect was not significantly changed by DPCPX, adenosine deaminase, yohimbine, bicuculline, 2-hydroxysaclofen and L-NAME (except that L-NAME enhanced the effect of kainate). AP5 and CNQX antagonized the immediate releasing effects in the same way that they antagonized the inhibition by AMPA, kainate and NMDA of the electrically evoked overflow of tritium.6. It is concluded that AMPA, kainate and NMDA, like glutamate, reduce the electrically evoked release of noradrenaline by releasing adenosine or an adenine nucleotide which is then degraded to adenosine. Activation of each of the three ionotropic glutamate receptors, AMPA, kainate and NMDA receptors, but not activation of metabotropic glutamate receptors can initiate this indirect inhibitory effect on the release of noradrenaline (as well as the known noradrenaline releasing effect).
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PMID:Ionotropic glutamate receptor types leading to adenosine-mediated inhibition of electrically evoked [3H]-noradrenaline release in rabbit brain cortex slices. 750 27

In anesthetized, paralyzed, and ventilated rats, hypoxia or intracarotid cyanide excited the carotid chemoafferents, whereas intracarotid dopamine and tyramine inhibited the chemoafferent discharges. The inhibition was abolished by chlorpromazine without attenuating the hypoxic excitation. Comparably, the hypoxic excitation was not attenuated by the following: 1) inhibition of nitric oxide synthase with NG-nitro-L-arginine; 2) inhibition of heme oxygenase with zinc protoporphyrin IX; 3) antagonism of ATP receptors with reactive blue 2; 4) antagonism of cholinergic receptors with atropine or trimethaphan; 5) inactivation of adenosine with adenosine deaminase; and 6) blockade of glutamate receptors with kynurenate. Systemic administration of ethylene glycol-bis(beta-aminoethyl ether)-N,N,N'N'-tetraacetic acid, in doses reversibly blocking sympathetic ganglionic transmission, was also without effect. Cyanide microinjection (0.05-0.5 nmol) into the petrosal but not nodose ganglion elicited a rapid dose-dependent elevation of arterial pressure. We conclude that excitation of the chemoreceptor afferents by hypoxia/cyanide cannot be attributed to release of these agents nor to others by Ca(2+)-dependent mechanisms. The results suggest that the afferent nerves themselves might function as oxygen detectors.
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PMID:Dopamine or transmitter release from rat carotid body may not be essential to hypoxic chemoreception. 752 4

A double-copy Moloney murine leukemia virus-based retroviral construct containing both the NEOr gene and a mutated dihydrofolate reductase cDNA (Leu 22-->Arg) was used to infect mouse bone marrow cells. The infected mouse marrow was returned to lethally irradiated mice. Primary, secondary, and even tertiary recipients transplanted with bone marrow cells infected with the recombinant virus showed protection from lethal methotrexate toxicity. The viral construct containing a SV-40 promoter in the U3 region of the 3' long terminal repeat appeared to be more effective than a similar construct containing the adenosine deaminase promoter, although both afforded protection. Evidence for integration into blood cells of both the NEOr gene and the mutated dihydrofolate reductase gene was obtained by polymerase chain reaction; sequencing of the amplified dihydrofolate reductase cDNA showed the presence of the point mutation. These results indicate that early hematopoietic progenitor cells in the mouse can be successfully transduced with a drug resistance gene.
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PMID:Long-term protection of recipient mice from lethal doses of methotrexate by marrow infected with a double-copy vector retrovirus containing a mutant dihydrofolate reductase. 762 Dec 35

The pyridoxal phosphate-dependent enzyme 1-aminocyclopropane-1-carboxylate synthase (ACC synthase; S-adenosyl-L-methionine methylthioadenosine-lyase, EC 4.4.1.14) catalyzes the conversion of S-adenosylmethionine (AdoMet) to ACC and 5'-methylthioadenosine, the committed step in ethylene biosynthesis in plants. Apple ACC synthase was overexpressed in Escherichia coli (3 mg/liter) and purified to near homogeneity. A continuous assay was developed by coupling the ACC synthase reaction to the deamination of 5'-methylthioadenosine by adenosine deaminase (adenosine aminohydrolase, EC 3.5.4.4) from Aspergillus oryzae. The enzyme is dimeric, with kcat = 9s-1 per monomer and Km = 12 microM for AdoMet. The pyridoxal phosphate-binding site of ACC synthase appears to be highly homologous to that of aspartate aminotransferase, suggesting similar roles for corresponding residues. Site-directed mutagenesis of Lys-273, Arg-407, and Tyr-233 (corresponding to residues 258, 386, and 225 in aspartate aminotransferase) and kinetic analyses of the mutants confirms their importance in the ACC synthase mechanism. The Lys-273 to Ala mutant has no detectable activity, supporting the identification of this residue as the base catalyzing C alpha proton abstraction. Mutation of Arg-407 to Lys results in a precipitous drop in kcat/Km and an increase in Km for AdoMet of at least 20-fold, in accordance with its proposed role as principal ligand for the substrate alpha-carboxylate group. Replacement of Tyr-233 with Phe causes a 24-fold increase in the Km for AdoMet and no change in kcat, suggesting that this residue plays a role in orienting the pyridoxal phosphate cofactor in the active site.
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PMID:Expression of apple 1-aminocyclopropane-1-carboxylate synthase in Escherichia coli: kinetic characterization of wild-type and active-site mutant forms. 780 54

We studied the role and relationship of the putative mediators of coupling of cerebral blood flow (CBF) and neuronal activation, adenosine (Ado) and nitric oxide (NO). Topical brain application over the whisker barrel cortex of anesthetized rats (n = 24) of the Ado receptor antagonist theophylline (Theo, 5 x 10(-5) M) for 30 min reduced the CBF response to deflection of the contralateral whiskers from 17.9 +/- 3.0% of baseline to 10.6 +/- 2.7% (P < 0.05). Coapplication of Theo (5 x 10(-5) M) and the NO synthase blocker N omega-nitro-L-arginine (L-NNA, 10(-3) M) for 30 min led to a further reduction in the CBF response to whisker stimulation to 7.5 +/- 1.3% (P < 0.05 compared with Theo alone). The CBF effect of sodium nitroprusside (10(-5) M) was not affected by Theo-L-NNA coapplication (122 +/- 25 vs. 140 +/- 25%, n = 5). Application of adenosine deaminase (1 U/ml, n = 5) reduced the CBF response to whisker stimulation from 18.2 +/- 0.7 to 10.7 +/- 1.9% (P < 0.05). Superfusion of L-NNA (10(-3) M, 30 min, n = 7) attenuated the CBF response to application of Ado (10(-4) M) from 39.4 +/- 10.4 to 22.9 +/- 10.5% (P < 0.05). N omega-nitro-D-arginine did not affect the CBF response to Ado (n = 5). We conclude that 1) Ado is involved in coupling of CBF to neuronal activation, 2) NO is involved in this response as well, and 3) there is an interaction between the vasodilator pathways of Ado and NO.
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PMID:Coupling of cerebral blood flow to neuronal activation: role of adenosine and nitric oxide. 804 94

Genetic deficiency of adenosine deaminase (ADA) results in varying degrees of immunodeficiency, including neonatal onset severe combined immunodeficiency (ADA- SCID) and milder, later onset immunodeficiency. We have determined the molecular basis of disease in a child from a consanguineous mating with ADA- SCID of clinically and biochemically reduced severity, diagnosed at 15 months of age and characterized by retention of more immunologic function than is typical of the fulminant neonatal onset type. The course was notable for an early predominance of bacterial infections and eosinophilia. In contrast to its absence in most ADA- SCIDs, residual ADA activity (1-2% of normal) could be detected in EBV-transformed B cells. Consistent with the increased residual ADA, excretion of the substrate deoxyadenosine and accumulation of the toxic metabolite deoxyATP were less than seen in ADA- SCID patients with fulminant disease. Sequence analysis of cDNA revealed a G853C transversion, predicting a substitution of proline for arginine at codon 253 (Arg253Pro). The parents were heterozygous and the child was homozygous for the mutation, as shown by sequence analysis of amplified genomic DNA. Transient expression of mutant cDNA in Cos cells revealed an electrophoretically abnormal, more negatively charged ADA with 1-2% of normal activity. These observations are consistent with replacement of positively charged arginine by proline, the lower accumulation of toxic metabolites, and the milder phenotype. By contrast, transient expression of a Gly216Arg mutant cDNA, associated, when homozygous, with neonatal onset ADA-SCID, did not reveal ADA activity. Mutations such as Arg253Pro, which retain residual activity of monomeric ADA, should be dominant for ameliorating the phenotype in patients carrying two different allelic mutations. Identification of additional similar mutations may be significant in evaluating the goals for and efficacy of current trials of gene and gene product replacement.
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PMID:Severe combined immunodeficiency of reduced severity due to homozygosity for an adenosine deaminase missense mutation (Arg253Pro). 825 46

Mutations at the adenosine deaminase (ADA) locus can result in varying degrees of immunodeficiency, including rapidly fulminant severe combined immunodeficiency (SCID) as well as a slowly progressive immunodeficiency not diagnosed until later in childhood. Genetic heterogeneity is a factor in the clinical heterogeneity. We have now identified, by direct sequencing of PCR-amplified genomic DNA, a G to A transition at a CpG dinucleotide predicting a glycine to arginine substitution at codon 20 (G20R). The mutation, in homozygosity, was associated with neonatal-onset rapidly fatal SCID. Consistent with homozygosity, the child was derived from a small isolated inbred community in Newfoundland. The mutation abolishes a site for the restriction enzyme BamHI and can be simply detected by agarose gel electrophoresis following amplification of exon 2 from genomic DNA and digestion with BamHI. The majority of ADA missense mutations can now be detected by similar amplification and enzyme digestion. We demonstrated that the G20R mutation is deleterious since introduction of the mutation into a normal ADA minigene abolished enzyme activity, as determined by transient expression in monkey kidney (Cos) cells. The amino acid substitution occurs in an area of the molecule conserved from Escherichia coli to man and that, as shown by crystallographic analysis, is involved in the binding of Zn2+ at the catalytic site. Although the mutation is in a CpG dinucleotide, known "hotspots" for G to A transitions, it was not found in a series of 43 additional ADA- chromosomes. Identification of mutations in additional ADA- patients with immunodeficiency of varying severity should further define the role that genotype plays in determining the extent of immunologic dysfunction.
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PMID:Homozygosity for a missense mutation (G20R) associated with neonatal onset adenosine deaminase-deficient severe combined immunodeficiency (ADA-SCID). 829 33

In the present study the effect of adenosine and adenosine analogues on rabbit isolated cavernosal smooth muscle has been evaluated in comparison with the effect of acetylcholine and electrical field stimulation. In the presence of guanethidine and indomethacin, acetylcholine and electrical field stimulation relaxed the rabbit corpus cavernosum, which was precontracted with phenylephrine. The nitric oxide synthesis inhibitor, N omega-nitro-L-arginine-methylester (L-NAME), greatly reduced the relaxation induced by electrical stimulation and completely abolished the relaxant effect of acetylcholine. A concentration-dependent relaxation of the rabbit corpus cavernosum was produced by adenosine; this effect was not modified by L-NAME, but was reduced by adenosine deaminase. On the other hand, the adenosine-induced relaxation was potentiated by the inhibitor of adenosine deaminase, erythro-9-(2-hydroxy-3-nonyl)adenine and by the adenosine uptake inhibitor dipyridamole. Moreover, the effect of adenosine was antagonized by the unspecific adenosine receptor antagonist 8-phenyltheophylline. The receptor subtypes involved in cavernosal relaxation were characterized by using selective receptor antagonists: 1,3-dipropyl-8-cyclopentylxanthine, a blocker of A, receptors, did not modify adenosine-induced relaxation. This effect was, however, antagonized by the A2-receptor antagonist CGS15943. A relaxant effect was also obtained with nanomolar concentrations of two synthetic adenosine analogues, the preferential A2 receptor agonist 5'-N-ethylcarboxamidoadenosine and the A2a selective agonist CGS21680. These results demonstrated that adenosine has potent relaxant activity on the corpus cavernosum, acting through a mechanism different from the nitric oxide pathway, and that receptors involved in the effect of adenosine belong to the A2a subtype.
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PMID:The potent relaxant effect of adenosine in rabbit corpora cavernosa is nitric oxide independent and mediated by A2 receptors. 853 48

Double-stranded RNA (dsRNA)-specific adenosine deaminase (DRADA) has been implicated as an enzyme responsible for the editing of RNA transcripts encoding glutamate-gated ion channel subunits (GLuR) in brain. In one case, the editing alters the gene-encoded glutamine (Q) to an arginine (R) located within the channel-forming domain of the alpha-amino-3-hydroxy-5-methyl-isoxazole-4-propionic acid (AMPA) receptor subunit GLuR-B. The result of editing at this site, called the 'Q/R' site, is a profound alteration of the Ca2+ permeability of the GLuR channel. Using recombinantly expressed DRADA proteins, we now demonstrate in vitro that DRADA is indeed involved in editing of the GLuR-B RNA. In addition to the formation of an RNA duplex structure involving exon and intron sequences, Q/R site-selective editing by DRADA also requires a cofactor protein(s) commonly present even in non-neuronal cells. The accuracy and efficiency of this RNA editing system appear to be determined by the quantitative balance between DRADA, cofactor and substrate GLuR-B RNA.
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PMID:Editing of the GLuR-B ion channel RNA in vitro by recombinant double-stranded RNA adenosine deaminase. 859 4

His 238, a conserved amino acid located in hydrogen-bonding distance from C-6 of the substrate in the active site of murine adenosine deaminase (mADA) and postulated to play an important role in catalysis, was altered into an alanine, a glutamate, and an arginine using site-directed mutagenesis. The Ala and Glu substitutions did not result in changes of the secondary or tertiary structure, while the Arg mutation caused local perturbations in tertiary structure and quenched the emission of one or more enzyme tryptophans. Neither the Glu or Arg mutations affected substrate binding affinity. By contrast, the Ala mutation enhanced substrate and inhibitor binding by 20-fold. The most inactive of the mutants, Glu 238, had a kcat/K(m) 4 x 10(-6) lower than the wild-type value, suggesting that a positive charge on His 238 is important for proper catalytic function. The Ala 238 mutant was the most active ADA, with a kcat/K(m) 2 x 10(-3) lower than the wild-type value. NMR spectroscopy and crystallography revealed that this mutant is able to catalyze hydration of purine riboside, a ground-state analog of the reaction. These results collectively show that His 238 is not required for formation of the hydroxylate used in the deamination and may instead have an important electrostatic role.
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PMID:Site-directed mutagenesis of histidine 238 in mouse adenosine deaminase: substitution of histidine 238 does not impede hydroxylate formation. 894 68


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