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:3.5.1.4 (deaminase)
5,113 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Trypanothione, the essential metabolite in the oxidant defense system of trypanosomatids, is synthesized by two distinct proteins, glutathionylspermidine synthetase and trypanothione synthetase. Glutathionylspermidine synthetase was purified to homogeneity from the trypanosomatid Crithidia fasciculata by aqueous two-phase systems and chromatography. The enzyme showed a specific activity of 38 micromol of glutathionylspermidine formed per min per mg of protein. Its molecular mass was 78 kDa in SDS-polyacrylamide gel electrophoresis, and it appeared predominantly monomeric in native polyacrylamide gel electrophoresis and gel filtration. The isoelectric point was at pH 4.6, and the pH optimum was near 7.6. Partial amino acid sequencing revealed homology with, but low similarity to, the glutathionylspermidine synthetase/amidase of Escherichia coli, and amidase activity was not detected in glutathionylspermidine synthetase of C. fasciculata. The kinetics of trypanosomatid glutathionylspermidine synthetase revealed a rapid equilibrium random mechanism with limiting Km values for Mg2+-ATP, GSH, and spermidine of 0.25 +/- 0.02, 2.51 +/- 0.33, and 0.47 +/- 0. 09 mM, respectively, and a kcat of 415 +/- 78 min-1. Partial reactions at restricted cosubstrate supply were not detected by 31P NMR, supporting the necessity of a quarternary complex formation for catalysis. ADP inhibited competitively with respect to ATP (Ki = 0. 08 mM) and trypanothione exerted a feedback inhibition competitive with GSH (Ki = 0.48 mM).
...
PMID:Convenient isolation and kinetic mechanism of glutathionylspermidine synthetase from Crithidia fasciculata. 911 52

A novel and effective treatment of biological samples with a combination of adenosine phosphate deaminase and apyrase was developed for reducing extracellular ATP, which has been a major problem encountered in improving the sensitivity of assays for intracellular ATP by the firefly luciferin-luciferase (L-L) method. Under the enzymatic reaction conditions, ATP and the related adenosine derivatives were converted to IMP, which are not active to the L-L system. In the model system (3.2 x 10(-8) M ATP in 1% yeast extract solution) the treatment with adenosine phosphate deaminase resulted in the reduction of ATP to 1.3 x 10(-11) M, and the concomitant use of apyrase lowered the concentration to 3.3 x 10(-13) M. The treatment (0.05 U/ml of adenosine phosphate deaminase and apyrase) was applied to the detection of bacteria in broth by the L-L method, affording the detection of 42 colony-forming unit (CFU)/ml of Escherichia coli and 10 CFU/ml of Staphylococcus aureus in the broth.
...
PMID:Enzymatic treatment to eliminate the extracellular ATP for improving the detectability of bacterial intracellular ATP. 924 33

Glutathionylspermidine (Gsp) is a metabolite common to Escherichia coli and protozoal parasites of the Trypanosoma family. Though its role in E. coli is unknown, Gsp is known to be an intermediate in the biosynthesis of N1,N8-bis(glutathionyl)spermidine (trypanothione), a metabolite unique to trypanosomatids that may allow the parasites to overcome oxidative stresses induced by host defense mechanisms. The bifunctional Gsp-synthetase/amidase from E. coli catalyzes both amide bond formation and breakdown between the N1-amine of spermidine [N-(3-aminopropyl)-1,4-diaminobutane] and the glycine carboxylate of glutathione (gamma-Glu-Cys-Gly), with net hydrolysis of ATP [Bollinger et al. (1995) J. Biol. Chem. 270 (23), 14031-14041]. Synthetase and amidase activities reside in separate domains of the protein, and liberation of the amidase domain from the synthetase domain activates the amidase activity as much as 70-fold in kcat/K(m) for a chromogenic substrate gamma-Glu-Ala-Gly-pNA [Kwon et al., (1997) J. Biol. Chem. 272 (4), 2429-2436]. When substrates for the Gsp-synthetase activity are present (GSH, ATP-Mg2+), Gsp-amidase is highly activated (15-fold). We provide kinetic and mutagenesis evidence suggesting that the amidase operates by a nucleophilic attack mechanism involving cysteine as the catalytic nucleophile. Stopped-flow studies on the 25 kDa Gsp-amidase fragment and the 70 kDa full-length Gsp-synthetase/amidase with gamma-Glu-Ala-Gly-ONp demonstrate burst kinetics characteristic of a covalent acyl-enzyme intermediate. Studies using various group-specific protease inhibitors, such as iodoacetamide, suggest an active-site cysteine or histidine as being relevant to amidase activity, and site-directed mutagenesis indicates that Cys-59 is essential for amidase activity.
...
PMID:Evidence for a glutathionyl-enzyme intermediate in the amidase activity of the bifunctional glutathionylspermidine synthetase/amidase from Escherichia coli. 939 17

Antimicrobial susceptibility testing by the ATP-bioluminescence method has been noted for its speed; it provides susceptibility results within 2 to 5 h. However, several disagreements between the ATP method and standard methodology have been reported. The present paper describes a novel ATP method in a 3.5-h test which overcomes these deficiencies through the elimination of false-resistance discrepancies in tests on gram-negative bacteria with beta-lactam agents. In our test model using Pseudomonas aeruginosa and piperacillin, it was shown that ATP in filamentous cells accounted for the false resistance. We found that 0.5% 2-amino-2-methyl-1,3-propanediol (AMPD) extracted ATP from the filamentous cells without affecting normal cells and that 0.3 U of adenosine phosphate deaminase (APDase)/ml simultaneously digested the extracted ATP. We used the mixture of these reagents for the pretreatment of cells in a procedure we named filamentous cell treatment, prior to ATP measurements. This novel ATP method with the filamentous cell treatment eliminated false-resistance discrepancies in tests on P. aeruginosa with beta-lactam agents, including piperacillin, cefoperazone, aztreonam, imipenem-cilastatin, ceftazidime, and cefsulodin. Furthermore, this novel methodology produced results which agreed with those of the standard microdilution method in other tests on gram-negative and gram-positive bacteria, including P. aeruginosa, Escherichia coli, Staphylococcus aureus, and Enterococcus faecalis, for non-beta-lactam agents, such as fosfomycin, ofloxacin, minocycline, and aminoglycosides. MICs obtained by the novel ATP method were also in agreement with those obtained by the agar dilution method of susceptibility testing. From these results, it was shown that the novel ATP method could be used successfully to test the activities of antimicrobial agents with the elimination of the previously reported discrepancies.
...
PMID:Novel antibiotic susceptibility tests by the ATP-bioluminescence method using filamentous cell treatment. 962 85

We have blocked creatine kinase (CK) mediated phosphocreatine (PCr) <==> ATP transphosphorylation in mitochondria and cytosol of skeletal muscle by knocking out the genes for the mitochondrial (ScCKmit) and the cytosolic (M-CK) CK isoforms in mice. Animals which carry single or double mutations, if kept and tested under standard laboratory conditions, have surprisingly mild changes in muscle physiology. Strenuous ex vivo conditions were necessary to reveal that MM-CK absence in single and double mutants leads to a partial loss of tetanic force output. Single ScCKmit deficiency has no noticeable effects but in combination the mutations cause slowing of the relaxation rate. Importantly, our studies revealed that there is metabolic and cytoarchitectural adaptation to CK defects in energy metabolism. The effects involve mutation type-dependent alterations in the levels of AMP, IMP, glycogen and phosphomonoesters, changes in activity of metabolic enzymes like AMP-deaminase, alterations in mitochondrial volume and contractile protein (MHC isoform) profiles, and a hyperproliferation of the terminal cysternae of the SR (in tubular aggregates). This suggests that there is a compensatory resiliency of loss-of-function and redirection of flux distributions in the metabolic network for cellular energy in our mutants.
...
PMID:Cytoarchitectural and metabolic adaptations in muscles with mitochondrial and cytosolic creatine kinase deficiencies. 974 21

The secondary structure of pig heart AMP-deaminase (AMP-d) in the absence and in the presence of orthophosphate or dioleoyl phosphatidic acid (DOPA) or ATP was investigated by FT-IR spectroscopy. While the latter substance activates the enzyme, orthophosphate is a well-known negative allosteric effector and DOPA exerts a noncompetitive inhibition on AMP-deaminase. Small changes in the secondary structure of AMP-d were induced by the above mentioned substances. Only DOPA reduced the thermal stability of AMP-d and avoided protein intermolecular interactions suggesting structural-functional relationships in AMP-d in the presence of the above substances and a possible role of phosphatidic acid in the subtle regulation of AMP-d activity by temporary binding of the enzyme to cellular membranes.
...
PMID:Structural-functional relationships in pig heart AMP-deaminase in the presence of ATP, orthophosphate, and phosphatidate bilayers. 978 95

A novel bioluminescent enzymatic cycling assay for ATP and AMP with concomitant use of firefly luciferase and pyruvate orthophosphate dikinase (PPDK) was developed. In this system, AMP and pyrophosphate produced from ATP by firefly luciferase were converted back into ATP by PPDK. This resulted in constant luminescence once the stable phase had been reached. Background luminescence of the reagent was reduced with adenosine phosphate deaminase by degrading ATP and AMP in the reagent. The maximum recycling ratio calculated from the integrated luminescence value was 2.64 cycles/min. The measurable ranges for ATP and AMP were equal and were between 4 x 10(-13) and 4 x 10(-17) mol/assay. The amount of yeast RNA could be estimated in the range of 1 x 10(-8) to 1 x 10(-12) g/assay by estimating the amount of AMP resulting from the degradation of RNA with nuclease P1. Various food samples were subjected to measurement of the amount of ATP + AMP + RNA to provide an index for hygiene monitoring. For beef extract, sensitivity was improved by more than 20 million compared to the previous methods relying only on the amount of ATP as an index.
...
PMID:An enzymatic cycling method using pyruvate orthophosphate dikinase and firefly luciferase for the simultaneous determination of ATP and AMP (RNA). 1003 67

We recently demonstrated that conditioned medium (CM) from peritoneal macrophages or activated microglia triggers a predominantly apoptotic death in hippocampal neurons in culture. We tested the effects of propentofylline (ppf), an agent that is neuroprotective in focal ischemia and is also associated with reduced microglial antigen expression after insult. Ppf had no impact on the secretion of neurotoxin from microglia. However, ppf significantly attenuated the effects of macrophage and microglial conditioned medium on neurons. Ppf did not attenuate neuronal hypoxic injury but did reverse the exaggeration of hypoxic injury exerted by subsequent addition of macrophage CM. A1 and A2 adenosine receptor inhibitors and an inhibitor of adenosine uptake each mimicked the effect of ppf. Neither ATP nor a deaminase inhibitor blocked the effect of microglial CM. These findings may be relevant to the neuroprotective effects of ppf in ischemia and dementia.
...
PMID:Propentofylline protects neurons in culture from death triggered by macrophage or microglial secretory products. 1021 75

Adenosine monophosphate deaminase (AMPD; EC 3.5.4.6) catalyses the hydrolysis of adenosine monophosphate (AMP) to commensurate amounts of inosine monophosphate (IMP) and ammonia. The production of AMP deaminase in Candida albicans was measured in Lee's medium grown cultures. The highest AMPD activity was observed at 24 h of growth. The enzyme had an optimum pH and temperature at 6-7 and 28 degrees C, respectively. This enzyme was inhibited under iron-limited growth conditions as well as by protease inhibitors. The AMPD of C. albicans showed a moderate increase in activity when cultures were grown in the presence of the divalent cations Mg2+, Ca2+, and Zn2+. Moreover, ADP, ATP, adenine, adenosine, deoxyribose and hypoxanthine increased the enzyme activity. Cultures grown in trypticase soy broth exhibited maximum AMPD activity compared with those grown in Sabouraud dextrose broth or Lee's medium.
...
PMID:Properties of adenosine monophosphate deaminase of Candida albicans. 1039 42

Extracellular adenosine (Ado) and ATP stimulate astrocyte proliferation through activation of P(1) and P(2) purinoceptors. Extracellular GTP and guanosine (Guo), however, that do not bind strongly to these receptors, are more effective mitogens than ATP and Ado. Exogenous Guo, like GTP and 5'-guanosine-betagamma-imidotriphosphate (GMP-PNP), dose-dependently stimulated proliferation of rat cultured astrocytes; potency order GMP-PNP > GTP > or = Guo. The mitogenic effect of Guo was independent of the extracellular breakdown of GTP to Guo, because GMP-PNP, a GTP analogue resistant to hydrolysis, was the most mitogenic. In addition to a direct effect on astrocytes, Guo exerts its proliferative activity involving Ado. Exogenous Guo, indeed, enhanced the extracellular levels of endogenous Ado assayed by HPLC in the medium of cultured astrocytes. Culture pretreatment with Ado deaminase (ADA), that converts Ado into inosine, reduced but did not abolish Guo-induced astrocyte proliferation whereas erythro-9-(2-hydroxy-3-nonyl)adenine (EHNA), that inhibits ADA activity, amplified Guo effect. Moreover, the mitogenic activity of Guo was partly inhibited by 8-cyclopentyl-1,3-dipropylxanthine and alloxazine, antagonists of Ado A(1) and A(2B) receptors, respectively. Also microglia seem to be a target for the action of Guo. Indeed, the mitogenic effect of Guo on astrocytes was: i) increased proportionally to the number of microglial cells present in the astrocyte cultures; ii) amplified when purified cultures of astrocytes were supplemented with conditioned medium deriving from Guo-pretreated microglial cultures. These data indicate that the mitogenic effects exerted by exogenous Guo on rat astrocytes are mediated via complex mechanisms involving extracellular Ado and microglia-derived soluble factors.
...
PMID:Cultured astrocyte proliferation induced by extracellular guanosine involves endogenous adenosine and is raised by the co-presence of microglia. 1064 47


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>

---