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)

Dihydroorotase +4,5-L-dihydro-orotate amidohydrolase [EC 3.5.2.3]), which catalyzes the reversible cyclization of N-carbamyl-L-aspartate to L-dihydroorotate, has been purified from orotate-grown Clostridium oroticum. The enzyme is homogeneous when subjected to polyacrylamide gel electrophoresis and is stable at pH 7.6 in 0.3 M NaCl containing 10 muM ZnSO4. The enzyme has a molecular weight of approximately 110,000. Sodium dodecyl sulfate gel electrophoresis, using three different buffer systems, indicated the enzyme is composed of two subunits, each having a molecular weight of 55,000. Dihydroorotase is shown by atomic absorption spectroscopy to be a zinc-containing metalloenzyme with 4 g-atoms of zinc per 110,000 g of protein. The pH optima for the conversion of N-carbamyl-L-aspartate to L-dihydroorotate and for L-dihydroorotate to N-carbamyl-L-aspartate are pH 6.0 and 8.2, respectively. The Km values for N-carbamyl-L-aspartate and for L-dihydroorotate are 0.13 and 0.07 mM, respectively. Inhibitor studies indicate that zinc may be involved in the catalytic activity of the enzyme.
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PMID:Purification of properties of dihydroorotase, a zinc-containing metalloenzyme in Clostridium oroticum. 0 24

Portions of closed jejunal biopsies from the dog were homogenised and their organelles separated by isopycnic centrifugation on continuous sucrose density gradients. The distributions of marker enzymes for the principal organelles were determined using highly sensitive assay procedures. The following organelles, with assayed marker enzymes and modal densities between brackets were characterised: peroxisomes (catalase, 1.21); brush borders (zinc-resistant alpha-glucosidase, leucyl-beta-naphthyl-amidase, gamma-glutamyl transferase, alkaline phosphatase, 1.20); lysosomes (N-acetyl-beta-glucosaminidase, alpha-mannosidase, 1.19); mitochondria (malate dehydrogenase, 1.18); endoplasmic reticulum (Tris-resistant alpha-glucosidase, 1.16); basal-lateral membranes (5'-nucleotidase, 1.11) and cytosol (lactate dehydrogenase). Homogenisation in isotonic sucrose containing digitonin (0.12 mmol/litre) selectively disrupted lysosomes and increased the equilibrium density of brush border and basal-lateral membranes. This procedure will be used to study the subcellular pathology of naturally occurring intestinal disease in the dog.
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PMID:Subcellular fractionation studies on peroral jejunal biopsies from the dog. 3 Jan 25

1. AMP-deaminase (EC 3.5.4.6) from skeletal muscle of frog and pikeperch was purified to homogeneity and compared with the homogeneous enzymes purified from rat, rabbit and hen skeletal muscle. 2. Their molecular weight was close to 280,000, every enzyme consisted of four identical subunits of molecular weight about 70,000. 3. All enzymes were found to contain about two atoms of zinc per molecule. 4. Minor differences of u.v.-absorption spectra between amphibian and fish muscle enzyme as compared with mammalian and bird muscle enzyme were found.
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PMID:Comparative studies on muscle AMP-deaminase--I. Purification, molecular weight, subunit structure and metal content of the enzymes from rat, rabbit, hen, frog and pikeperch. 4 53

The quantitative method for determination of penicillinacylase activity is described. The method is based on detection of phenylacetic acid (PAA) formed during hydrolysis of benzylpenicillin. PAA is extracted with toluol and nitrated with potassium nitrate solution in concentrated sulphuric acid followed by reduction of nitrophenylacetic acid into aminophenylacetic acid with zinc powder. Aminophenylacetic acid interacts with p-dimethylaminobenzaldehyde in acid medium forming a coloured compound (Schiff base). The optic density of the latter was determined with spectrophotometer SF-16 at 430 nm. The method was used for determination of acylase in Yersinia species and some other bacteria.
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PMID:[Method for the quantitative determination of penicillin acylase activity by the formation of phenylacetic acid]. 79 10

Further properties of cow liver deaminase are reported. Highly purified deaminase migrated as a single badn on starch and polycrylamide gels electrophoresis. Molecular weight determinations by means of gel filtration on calibrated columns of Sephadex G-100, Sepharose 4 B and B 10-Gel P-100, gave values of 40 000 +/- 4 000. Data obtained suggest that cow liver deaminase exists as a single polypeptide chain. Heating partially purified preparations of deminase resulted in an enhancement of activity. Added cosynthetase to these fractions increased the percentage of uroporphyrinogen III formed but also diminished total uroporphyrinogens synthesis. The action of several compounds added to the system was studied. Thiol reagents and divalent metals as Hg 2+ , Pb2+, (d2+ and Zn2+ inhibited deaminase, indicating the presence of thiol groups essential for activity, probably involved in the cyclization step. Certain concentrations of sodium, potassium and magnesium salts enhanced activity. Several chelators tested were without effect on the deminase. Some dicarboxylic acids and 2-methoxy-5-nitrotropone inhibited the enzyme.
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PMID:Studies on cow liver porphobilinogen deaminase. 105 1

The method of measuring enzyme deactivation by monitoring necessary addition of fresh enzyme to keep a constant degree of conversion in a CSTR at constant [E] x tau, the product of concentration of active enzyme [E] and residence time tau, was successfully applied to acylase I from porcine kidney and Aspergillus oryzae fungus. Fungal enzyme was found to be more stable than kidney enzyme. Activation by both Co2+ and Zn2+ ions also yielded increased operational enzyme stability: Co2+ and Zn2+ are better stabilizers than activators. Mg2+ and Ca2+ are found to be neither activators nor stabilizers. Fungal acylase partially deactivated by exposition to a metal-free medium in the CSTR was reactivated by addition of Zn2+, demonstrating that loss of Zn2+ from the enzyme molecule is mainly responsible for deactivation in a continuous reactor.
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PMID:Operational stability of enzymes. Acylase-catalyzed resolution of N-acetyl amino acids to enantiomerically pure L-amino acids. 147 69

The structural gene that encodes cytidine deaminase (cdd) in Escherichia coli was cloned from Kohara phage lambda 365 (7F1), and its nucleotide sequence was determined. Plasmids harboring the gene complemented chromosomal cdd mutations, enhanced cytidine deaminase activity in cell extracts, and directed the synthesis of a protein identical in mass and N-terminal amino acid sequence with cytidine deaminase purified from wild-type bacteria. Metal analysis of the purified, plasmid-encoded deaminase indicated a single atom of tightly bound zinc per subunit. Earlier work has shown that bacterial cytidine deaminase and mammalian adenosine deaminase are remarkably alike in their mechanisms of action, in their free energies of interaction with analogue inhibitors resembling tetrahedral intermediates in nucleophilic substitution, and in their ability to discriminate between analogue inhibitors differing by a single hydroxyl group. In contrast to these shared catalytic similarities, the deduced amino acid sequence of E. coli cytidine deaminase (monomer MW 31,540) differs markedly from the mammalian adenosine deaminase sequence suggesting major differences in their tertiary structures. Nevertheless, cytidine deaminase and mammalian plus bacterial adenosine deaminases share a single region (TVHA) of sequence identity that is tentatively identified as part of the cytidine deaminase active site.
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PMID:Cloning and nucleotide sequence of the Escherichia coli cytidine deaminase (ccd) gene. 156 63

Mammalian DHOase (S-dihydroorotate amidohydrolase, EC 3.5.2.3) is part of a large multifunctional protein called CAD, which also has a carbamoyl-phosphate synthetase [carbon-dioxide: L-glutamine amido-ligase (ADP-forming, carbamate-phosphorylating), EC 6.3.5.5] and aspartate transcarbamoylase (carbamoyl-phosphate: L-aspartate carbamoyltransferase, EC 2.1.3.2) activities. We sequenced selected restriction fragments of a Syrian hamster CAD cDNA. The deduced amino acid sequence agreed with the sequence of tryptic peptides and the amino acid composition of the DHOase domain isolated by controlled proteolysis of CAD. Escherichia coli transformed with a recombinant plasmid containing the cDNA segment 5' to the aspartate transcarbamoylase coding region expressed a polypeptide recognized by DHOase domain-specific antibodies. Thus, the order of domains within the polypeptide is NH2-carbamoyl-phosphate synthetase-DHO-aspartate transcarbamoylase-COOH. The 334-residue DHOase domain has a molecular weight of 36,733 and a pI of 6.1. A fragment of CAD having DHOase activity that was isolated after trypsin digestion has extensions on both the NH2 (18 residues) and COOH (47-65 residues) termini of this core domain. Three of five conserved histidines are within short, highly conserved regions that may participate in zinc binding. Phylogenetic analysis clustered the monofunctional and fused DHOases separately. Although these families may have arisen by convergent evolution, we favor a model involving DHOase gene duplication and insertion into an ancestral bifunctional locus.
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PMID:Mammalian dihydroorotase: nucleotide sequence, peptide sequences, and evolution of the dihydroorotase domain of the multifunctional protein CAD. 196 94

Glucosamine-6-phosphate isomerase deaminase from Escherichia coli, a typical allosteric enzyme, becomes less cooperative and 50% inhibited when treated with zinc. This metal cation behaving as a tight-bound and slow partial inhibitor. Modification of a pair of vicinal reactive thiols with some sulfhydryl reagents mimics this effect. On the other hand, sulfhydryl reactivity disappears in the presence of saturating concentrations of Zn2+, which does not modify the kinetics of S-methylated enzyme, a finding that indicates that vicinal thiols are an essential part of the zinc-binding site. Allosteric activation of the deaminase causes trapping of the metal, which cannot be released by dialysis against a buffer containing EDTA. Cadmium and nickel(II) cations also produce a similar effect.
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PMID:Zinc binding and its trapping by allosteric transition in glucosamine-6-phosphate deaminase from Escherichia coli. 211 Nov 70

A recombinant trypsin was designed whose catalytic activity can be regulated by varying the concentration of Cu2+ in solution. Substitution of Arg-96 with a His in rat trypsin (trypsin R96H) places a new imidazole group on the surface of the enzyme near the essential active-site His-57. The unique spatial orientation of these His side chains results in the formation of a stable, metal-binding site that chelates divalent first-row transition-metal ions. Occupancy of this site by a metal ion prevents the imidazole group of His-57 from participating as a general base in catalysis. As a consequence, the primary effect of the transition metal ion is to inhibit the esterase and amidase activities of trypsin R96H. The apparent Ki for this inhibition is in the micromolar range for copper, nickel, and zinc, the tightest binding being to Cu2+ at 21 microM. Trypsin R96H activity can be fully restored by removing the bound Cu2+ ion with EDTA. Multiple cycles of inhibition by Cu2+ ions and reactivation by EDTA demonstrate that reversible regulatory control has been introduced into the enzyme. These results describe a novel mode of inhibition of serine protease activity that may also prove applicable to other proteins.
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PMID:Regulation of serine protease activity by an engineered metal switch. 212 68


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