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

Several commercially available enzymes were tested for their ability to hydrolyze amino acid carbamates. No activity was found with pig liver esterase, the hydantoinase from Pseudomonas fluorescens DSM 84, or the urease from jack beans. A stereoselective cleavage of the carbamyl group yielding L-amino acids was observed by acylase and acetylcholinesterases from bovine and human erythrocytes. Racemic mixtures of N-(methoxycarbonyl)-DL-alanine, N-(ethoxycarbonyl)-DL-alanine, and the corresponding valine carbamates are hydrolyzed to L-alanine and L-valine, respectively, by acylases leaving the D-amino acid carbamates unchanged. The lysine carbamates were not hydrolyzed by acylases. In contrast only the methoxycarbonyl amino acids were split by acetylcholinesterases, which, however, also cleave alpha, epsilon-(N-methoxycarbonyl)-DL-lysine stereoselectively at the alpha position, yielding epsilon-N-methoxycarbonyl-L-lysine. The optimum pH for enzymatic activity of hog kidney acylase was 7.5 and a Km value of 8.2 mM for N-(methoxycarbonyl)-DL-alanine was determined. For the acetylcholinesterases the reaction rate reaches an optimum between pH 7.5 and 8. The Km value was 68 mM for N-(methoxycarbonyl)-DL-alanine.
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PMID:Studies on the enzymatic hydrolysis of amino acid carbamates. 311 96

We have developed a one-dimensional thin-layer chromatography procedure that resolves the initial substrate uracil and its catabolic derivatives dihydrouracil, N-carbamoyl-beta-alanine (NCBA) and beta-alanine. This separation scheme also simplifies the preparation of the radioisotopes of N-carbamoyl-beta-alanine and dihydrouracil. Combined, these methods make it possible to assay easily and unambiguously, jointly or individually, all three enzyme activities of uracil catabolism: dihydropyrimidine dehydrogenase, dihydropyrimidinase, and N-carbamoyl-beta-alanine amidohydrolase. Earlier reports had presented data suggesting that these three enzyme activities were combined in a complex because they appeared to be controlled at a single genetic locus [Dagg, C. P., Coleman, D.L., & Fraser, G.M. (1964) Genetics 49, 979-989] and because they appeared able to channel metabolites [Barrett, H.W., Munavalli, S.N., & Newmark, P. (1964) Biochim. Biophys. Acta 91, 199-204]. Although the three enzymes from rat liver have similar sizes, with apparent molecular weights of 218 000 for dihydropyrimidine dehydrogenase, 226 000 for dihydropyrimidinase, and 234 000 for NC beta A amidohydrolase, they are easily separated from each other. Kinetic studies show no evidence of substrate channeling and therefore do not support a model for an enzyme complex. The earlier reports may be explained by our studies on the amidohydrolase, which suggest that under certain conditions this enzyme may become the rate-limiting step in uracil catabolism.
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PMID:Pyrimidine catabolism: individual characterization of the three sequential enzymes with a new assay. 643 73

Pyrimidine ribonucleoside catabolic enzyme activities of the opportunistic pathogen Pseudomonas pickettii were examined. Of the pyrimidine and related compounds tested, only dihydrouracil (nitrogen source) and ribose (carbon source) supported growth. Thin-layer chromatographic separation of the uridine and cytidine catabolities produced by P. pickettii extracts indicated that this pseudomonad contained nucleoside hydrolase activity. Its presence was confirmed by enzyme assay. Hydrolase activity was elevated in both glucose- and ribose-grown cells relative to succinate-grown cells. Nucleoside hydrolase activity was depressed when dihydrouracil served as a nitrogen source. Cytosine deaminase activity was present in extracts prepared from succinate-, glucose- or ribose-grown cells when (NH4)2SO4 served as the nitrogen source although cells grown on glucose or ribose exhibited a higher enzyme activity. Cytosine deaminase activity was not detected in extracts prepared from cells grown on dihydrouracil as a nitrogen source. Both dihydropyrimidine dehydrogenase and dihydropyrimidinase activities were measurable in P. pickettii. The dehydrogenase activity was higher with NADH than with NADPH as its nicotinamide cofactor when uracil served as its substrate. Carbon source did not affect dehydrogenase or dihydropyrimidinase activity greatly but both activities were diminished in cells grown on the nitrogen source dihydrouracil.
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PMID:Pyrimidine ribonucleoside catabolic enzyme activities of Pseudomonas pickettii. 771 Feb 77

A thermostable N-carbamoyl-D-amino acid amidohydrolase was found in the cells of newly isolated bacterium. Blastobacter sp. A17p-4. The bacterium also showed D-specific hydantoinase activity. The N-carbamoyl-D-amino acid amidohydrolase activity of the cells exhibited a temperature optimum at 50-55 degrees C, and was stable up to 50 degrees C. The N-carbamoyl-D-amino acid amidohydrolase of Blastobacter sp. A17p-4 was purified to homogeneity and characterized. It has a relative molecular weight of about 120,000 and consists of three identical subunits with a relative molecular weight of about 40,000. N-Carbamoyl-D-amino acids having hydrophobic groups served as good substrates for the enzyme. It has been suggested that D-amino acid production from DL-5-substituted hydantoin involves the action of a series of enzymes involved in pyrimidine degradation, namely amide-ring opening enzyme, dihydropyrimidinase, and N-carbamoylamide hydrolyzing enzyme, beta-ureidopropionase. However, the purified enzyme did not hydrolyze beta-ureidopropionate; suggesting that the N-carbamoyl-D-amino acid amidohydrolase coexisting with D-specific hydantoinase, probably dihydropyrimidinase, in Blastobacter sp. A17p-4 is different from beta-ureidopropionase.
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PMID:Thermostable N-carbamoyl-D-amino acid amidohydrolase: screening, purification and characterization. 776 78

The mechanism of stereospecific conversion of DL-5-substituted hydantoins to the corresponding L-amino acids by Pseudomonas sp. strain NS671 was studied. The results indicated that the hydantoinase catalyzed the hydrolysis reaction of both D- and L-5-(2-methylthioethyl)hydantoin, and that the hydrolysis of the L-enantiomer proceeded preferentially compared with that of the D-enantiomer. On the basis of these findings, the mechanism was speculated to be as follows: DL-5-substituted hydantoins are converted exclusively to the L-forms of the corresponding N-carbamyl-amino acids by the hydantoinase in combination with hydantoin racemase. The N-carbamyl-L-amino acids are then converted to L-amino acids by N-carbamyl-L-amino acid amidohydrolase.
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PMID:Mechanism of stereospecific conversion of DL-5-substituted hydantoins to the corresponding L-amino acids by Pseudomonas sp. strain NS671. 902 51

Agrobacterium sp. strain KNK712, which produced N-carbamyl-D-amino acid amidohydrolase (DCase) was isolated from soil. The bacterium had D-specific hydantoinase activity also. Both enzymes are suitable for use in the production of D-amino acids. The DCase gene from Agrobacterium sp. strain KNK712 was cloned into Escherichia coli. The cloned DNA fragment contained one open reading frame, predicted to encode a peptide of 304 amino acids, with a calculated molecular weight of 34,285. The DCase gene was overexpressed under the control of the lac promoter, and DCase accounted for 50% of the soluble protein in the cells. The enzyme was purified and some properties were investigated. Both the optimum pH and the pH that gave greatest stability were about pH 7.0. The optimum temperature was 65 degrees C, and the enzyme was stable at 55 degrees C. The enzyme had strict specificity toward N-carbamyl-D-amino acids, and was inhibited by thiol reagents, Cu2+, Hg2+, Ag+, and ammonia.
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PMID:Isolation of Agrobacterium sp. strain KNK712 that produces N-carbamyl-D-amino acid amidohydrolase, cloning of the gene for this enzyme, and properties of the enzyme. 964 17

We developed a fully enzymatic process employing D-hydantoinase and N-carbamoylase for the production of D-amino acid from 5'-monosubstituted hydantoin. For the comparison of the reaction systems using two sequential enzymes, D-hydantoinase of Bacillus stearothermophilus SD1 and N-carbamoyl-D-amino acid amidohydrolase (N-carbamoylase) of Agrobacterium tumefaciens NRRL B11291 were separately expressed in each host cell and coexpressed in the same host cell. A high level and constitutive expression of both enzymes in Escherichia coli in a soluble form was achieved using a promoter derived from B. stearothermophilus SD1. The expression levels of both enzymes ranged from 17% to 23% of the total soluble protein, depending on the expression system. In the case of employing separately expressed enzymes, the product yield of D-hydroxyphenylglycine from D,L-p-hydroxyphenylhydantoin and productivity were 71% and 2.57 mM/g-cell/h in 15 h, respectively. The accumulation of N-carbamoyl-D-hydroxyphenylglycine was significant over the reaction time. On the other hand, use of coexpressed enzymes resulted in 98% product yield of D-hydroxyphenylglycine in 15 h, minimizing the level of intermediates in the reaction mixture. The productivity of coexpressed whole cell reaction was estimated to be 6.47 mM/g-cell/h in 15 h. The coexpressed system was tested for an elevated concentration of D,L-p-hydroxyphenylhydantoin, and efficient production can be achieved.
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PMID:Production of D-amino acid using whole cells of recombinant Escherichia coli with separately and coexpressed D-hydantoinase and N-carbamoylase. 1093 29

A superfamily of cyclic amidohydrolases, including dihydropyrimidinase, allantoinase, hydantoinase, and dihydroorotase, all of which are involved in the metabolism of purine and pyrimidine rings, was recently proposed based on the rigidly conserved structural domains in identical positions of the related enzymes. With these conserved domains, two putative cyclic amidohydrolase genes from Escherichia coli, flanked by related genes, were identified and characterized. From the genome sequence of E. coli, the allB gene and a putative open reading frame, tentatively designated as a hyuA (for hydantoin-utilizing enzyme) gene, were predicted to express hydrolases. In contrast to allB, high-level expression of hyuA in E. coli of a single protein was unsuccessful even under various induction conditions. We expressed HyuA as a maltose binding protein fusion protein and AllB in its native form and then purified each of them by conventional procedures. allB was found to encode a tetrameric allantoinase (453 amino acids) which specifically hydrolyzes the purine metabolite allantoin to allantoic acid. Another open reading frame, hyuA, located near 64.4 min on the physical map and known as a UUG start, coded for D-stereospecific phenylhydantoinase (465 amino acids) which is a homotetramer. As a novel enzyme belonging to a cyclic amidohydrolase superfamily, E. coli phenylhydantoinase exhibited a distinct activity toward the hydantoin derivative with an aromatic side chain at the 5' position but did not readily hydrolyze the simple cyclic ureides. The deduced amino acid sequence of the novel phenylhydantoinase shared a significant homology (>45%) with those of allantoinase and dihydropyrimidinase, but its functional role still remains to be elucidated. Despite the unclear physiological function of HyuA, its presence, along with the allantoin-utilizing AllB, strongly suggested that the cyclic ureides might be utilized as nutrient sources in E. coli.
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PMID:Functional expression and characterization of the two cyclic amidohydrolase enzymes, allantoinase and a novel phenylhydantoinase, from Escherichia coli. 1109 64

The vertebrate CRMP (collapsin-response-mediator protein) gene family comprises at least four members. These CRMPs exhibit about 60% amino acid identity with vertebrate dihydropyrimidinase (DHP), an amidohydrolase involved in the pyrimidine degradation pathway. CRMP is also referred to as DRP (DHP-related protein), TOAD-64 (turned on after division, 64 kDa) and Ulip (Unc-33-like phosphoprotein). These vertebrate CRMPs are expressed mainly in early neuronal differentiation, which suggests that they play a role in neuronal development. In this study we isolated two cDNA clones from nematode C. elegans based on their sequence homology to vertebrate CRMPs and DHP. These two molecules, termed CeCRMP/DHP-1 and -2, turned out to be Ulip-B and -A, respectively, which were previously identified in the C. elegans genomic database by Byk et al. (1998). These newly isolated molecules were believed to represent a common ancestral state before the gene duplication between CRMPs and DHP. CeCRMP/DHP-1 and -2 protein retained all putative zinc-binding residues thought to be essential for the amidohydrolase activity of DHP and exhibited a weak amidohydrolase activity when 5-bromo-dihydrouracil was used as a substrate. Whole-mount in situ hybridization and expression analysis using GFP fusions revealed that CeCRMP/DHP-1 was transiently expressed in the hypodermis of C. elegans during the early larva stage. CeCRMP/DHP-1 was also expressed in a single nerve cell between the pharynx and ring neuropil. On the other hand, expression of CeCRMP/DHP-2 was observed in the body wall muscle throughout the lifespan of C. elegans. These results indicate that a major site of CeCRMP/DHP-1 and -2 expression is non-neuronal. Targeted gene disruption of CeCRMP/DHP-2 caused no particular difference in appearance or movement phenotype.
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PMID:Cloning and characterization of the Caenorhabditis elegans CeCRMP/DHP-1 and -2; common ancestors of CRMP and dihydropyrimidinase? 1116 13

An interesting phenomenon was observed that the existence of the intact cell membrane can enhance the D-amino acids production from D,L-5-substituted hydantoins by reacting with the whole cells of Agrobacterium radiobacter. Two intracellular enzymes were involved in the reaction process. The first enzyme D-hydantoinase converted hydantoins to carbamoyl derivatives which were further converted to D-amino acids by D-amidohydrolase. The amount of D-amino acids produced from hydantoins by the intact cells were 1.8-2.4 fold higher than the toluene treated cells. In addition, by using the intact cells the amount of D-amino acids produced from hydantoins was about 10 fold higher than that produced directly from carbamoyl derivatives. The relatively lower permeability of cell membrane to the reaction intermediate carbamoyl derivatives was confirmed by a simple mathematical model to be the main factor for the better performance of the intact cells for D-amino acid production. Besides, the low intracellular enzymes activities also contributed to the effect of intact cell membrane on enhancing the D-amino acid production.
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PMID:Effect of cell membrane of Agrobacterium radiobacter on enhancing D-amino acids production from racemic hydantoins. 1139 62


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