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)

The kinetic constants for hydrolysis and transfer (with hydroxylamine as the alternate acceptor) of the aliphatic amidase (acylamide amidohydrolase, EC 3.5.1.4) from Pseudomonas aeruginosa were determined for a variety of acetyl and propionyl derivatives. The results obtained were consistent with a ping-pong or substitution mechanism. Product inhibition, which was pH dependent, implicated an acyl-enzyme compound as a compulsory intermediate and indicated that ammonia combined additionally with the free enzyme in a dead-end manner. The uncompetitive activation of acetamide hydrolysis by hydroxylamine and the observation that the partitioning of products between acetic acid and acetohydroxamate was linearly dependent on the hydroxylamine concentration substantiated these conclusions and indicated that deacylation was at least partially rate limiting. With propionamide as the acyl donor apparently anomalous results, which included inequalities in certain kinetic constants and a hyperbolic dependence of the partition ratio on the hydroxylamine concentration, could be explained by postulating a compulsory isomerisation of the acyl-enzyme intermediate prior to the transfer reaction.
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PMID:Kinetic mechanism of the aliphatic amidase from Pseudomonas aeruginosa. 11 Mar 50

Porphobilinogen-deaminase from Saccharomyces cerevisiae has been isolated and partially purified 80- and 230-fold in the absence or presence of phenylmethylsulphonyl fluoride, respectively. Some properties of the isolated enzyme were studied. Porphyrin formation was linear with time and protein concentration. Optimum pH was about 7.5-7.8. Molecular mass of the protein was 30,000 +/- 3000 Dalton when the enzyme was purified in the presence of phenylmethylsulphonyl fluoride. A less active and unstable 20,000 Da molecular mass species was obtained when purification was performed in the absence of the protease inhibitor. Porphobilinogen-deaminase exhibited classical Michaelis-Menten kinetics. The apparent Km for uroporphyrinogen formation was 19 microM; Vmax was 3.6 nmol uroporphyrin/h and the Hill coefficient was n = 1. Also the action of several reagents on the activity was studied. Protective thiol agents had no effect. Heavy metals inhibited both porphyrin formation and porphobilinogen consumption, but known sulphydryl inactivating chemicals inhibit the former without modifying the latter. Ammonium ions had no effect on the activity while hydroxylamine completely inhibited both porphyrin formation and porphobilinogen consumption.
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PMID:Studies on porphobilinogen-deaminase from Saccharomyces cerevisiae. 181 12

The effect of chemical modification on the pseudocholinesterase and aryl acylamidase activities of purified human serum pseudocholinesterase was examined in the absence and presence of butyrylcholine iodide, the substrate of pseudocholinesterase. Modification by 2-hydroxy-5-nitrobenzyl bromide, N-bromosuccinimide, diethylpyrocarbonate and trinitrobenzenesulfonic acid caused a parallel inactivation of both pseudocholinesterase and aryl acylamidase activities that could be prevented by butyrylcholine iodide. With phenylglyoxal and 2,4-pentanedione as modifiers there was a selective activation of pseudocholinesterase alone with no effect on aryl acylamidase. This activation could be prevented by butyrylcholine iodide. N-Ethylmaleimide and p-hydroxy-mercuribenzoate when used for modification did not have any effect on the enzyme activities. The results suggested essential tryptophan, lysine and histidine residues at a common catalytic site for pseudocholinesterase and aryl acylamidase and an arginine residue (or residues) exclusively for pseudocholinesterase. The use of N-acetylimidazole, tetranitromethane and acetic anhydride as modifiers indicated a biphasic change in both pseudocholinesterase and aryl acylamidase activities. At low concentrations of the modifiers a stimulation in activities and at high concentrations an inactivation was observed. Butyrylcholine iodide or propionylcholine chloride selectively protected the inactivation phase without affecting the activation phase. Protection by the substrates at the inactivation phase resulted in not only a reversal of the enzyme inactivation but also an activation. Spectral studies and hydroxylamine treatment showed that tyrosine residues were modified during the activation phase. The results suggested that the modified tyrosine residues responsible for the activation were not involved in the active site of pseudocholinesterase or aryl acylamidase and that they were more amenable for modification in comparison to the residues responsible for inactivation. Two reversible inhibitors of pseudocholinesterase, namely ethopropazine and imipramine, were used as protectors during modification. Unlike the substrate butyrylcholine iodide, these inhibitors could not protect against the inactivation resulting from modification by 2-hydroxy-5-nitrobenzyl bromide, N-bromosuccinimide and trinitrobenzenesulfonic acid. But they could protect against the activation of pseudocholinesterase and aryl acylamidase by low concentrations of N-acetylimidazole and acetic anhydride thereby suggesting that the binding site of these inhibitors involves the non-active-site tyrosine residues.
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PMID:Chemical modification of the bifunctional human serum pseudocholinesterase. Effect on the pseudocholinesterase and aryl acylamidase activities. 286 42

A simpler method for purifying human red cell deaminase, using a mixture of n-butanol and chloroform, which denatures hemoglobin, followed by ammonium sulphate fractionation, heat treatment, Sephadex G-100 and DEAE-cellulose chromatography, yielding a 3400 fold purified enzyme is described. Some properties of purified deaminase were studied. The enzyme seems to have a strict requirement for oxygen, neither PBG consumption nor uroporphyrinogens formation were measured under anaerobiosis. Uroporphyrinogens formation was linear with both protein and time over a wide range of enzyme concentration and up to 2 h. The optimum pH was 7.4 and the mol. wt was 40,000 +/- 4000. The enzyme was heat-stable and increased its activity by heating. Ammonium and hydroxylamine ions inhibited the reaction. K+ and Na+ ions did not greatly affect activity, while most divalent cations tested significantly diminished uroporphyrinogen formation and to a lesser degree PBG consumption. Direct plots of velocity against PBG concentration were hyperbolic, however double-reciprocal plots were non-linear, Hill plots gave an n value of 2 and Eadie plots were bell-shaped, indicating the existence of weakly positive cooperative effect between 2 binding sites for PBG per molecule of deaminase.
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PMID:Human red cell porphobilinogen deaminase. A simpler method of purification and some unusual properties. 400 48

The activities of six bacteriophage T2r(+)-induced enzymes (thymidylate synthetase, deoxycytidylate deaminase, thymidylate kinase, deoxycytidylate hydroxymethylase, deoxycytidine pyrophosphatase, and dihydrofolate reductase) were measured after dilution of phage-infected Escherichia coli B from 8 x 10(8) to 2 x 10(8) cells per ml. The only enzyme activity altered was that of deoxycytidylate deaminase, which increased three- to fourfold. Conversely, the rapid concentration of cells from 2 x 10(8) to 8 x 10(8) per ml did not result in a reduction in deaminase activity. Although an enhancement in aeration reduced the response of deoxycytidylate deaminase to cellular dilution, the influence of potential metabolic inhibitors or activators could not be shown. The change in deoxycytidylate deaminase activity appeared to be associated with an altered translational event, since the increase could not be prevented by rifampin but was blocked effectively by chloramphenicol and hydroxylamine. In addition, antibody to the T2 phage-induced deoxycytidylate deaminase demonstrated that the increase in enzyme activity was associated with a corresponding increase in radioactive leucine incorporated into the enzyme antigen.
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PMID:Relationship between Escherichia coli B titer and the level of deoxycytidylate deaminase activity induced on bacteriophage T2r + infection. 433 61

4-N-hydroxy-cytidine was found to substitute for uridine as a pyrimidine supplement for the growth of Escherichia coli Bu(-). Measurement of the incorporation of 4-N-hydroxy-cytidine-2-(14)C into ribonucleic acid and deoxyribonucleic acid revealed that this compound was converted to cytidine or uridine before utilization. Two pathways for metabolism were considered: (i) the reduction of 4-N-hydroxy-cytidine to cytidine followed by deamination, (ii) the direct hydrolysis of hydroxylamine from 4-N-hydroxy-cytidine to yield uridine. A threefold increase in cytidine (deoxycytidine) deaminase (EC 3.5.4.5) activity, when the cells were grown on 4-N-hydroxy-cytidine, suggested the involvement of this enzyme. More direct proof was obtained by purifying the deaminase 185-fold and finding that it released hydroxylamine from 4-N-hydroxy-cytidine at one-fiftieth the rate at which ammonia was removed from cytidine. This result is consistent with the slower rate of growth of the Bu(-) cells on 4-N-hydroxy-cytidine than cytidine and suggests that the second pathway is the major route for utilization of this compound.
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PMID:Metabolism of 4-N-hydroxy-cytidine in Escherichia coli. 494 53

1. The penicillin acylase of Eschericha coli N.C.I.B. 8743 is a reversible enzyme. Reaction rates for the two directions have been determined. 2. Measurements of the rates of enzymic synthesis of penicillins from 6-aminopenicillanic acid and various carboxylic acids revealed that p-hydroxyphenylacetic acid was the best substrate, followed by phenylacetic, 2-thienylacetic, substituted phenylacetic, 3-hexenoic and n-hexanoic acids. 3. The rate of synthesis of penicillin improved when amides or N-acylglycines were used; alpha-aminobenzylpenicillin and phenoxymethylpenicillin were only synthesized when using these more energy-rich compounds. 4. Phenyl-acetylglycine was the best substrate for the synthesis of benzylpenicillin compared with other derivatives of phenylacetic acid. 5. The enzyme was specific for acyl-l-amino acids, benzylpenicillin being synthesized from phenylacetyl-l-alpha-aminophenylacetic acid but not from phenylacetyl-d-alpha-aminophenylacetic acid. 6. alpha-Phenoxyethylpenicillin was synthesized from 6-aminopenicillanic acid and alpha-phenoxypropionylthioglycollic acid non-enzymically, but the rate was faster in the presence of the enzyme. 7. The E. coli acylase catalysed the acylation of hydroxylamine by acids or amides to give hydroxamic acids, the phenylacetyl group being the most suitable acyl group. The enzyme also catalysed other acyl-group transfers.
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PMID:Penicillins and other acylamino compounds synthesized by the cell-bound penicillin acylase of Escherichia coli. 498 18

1. Porphobilinogenase was isolated and purified from soya-bean callus tissue; its components, porphobilinogen deaminase and uroporphyrinogen isomerase, were separated and purified. 2. The purified porphobilinogenase was resolved into two bands on starch-gel electrophoresis. The molecular weights of porphobilinogenase, deaminase and isomerase fractions were determined by the gel-filtration method. Porphobilinogenase activity was affected by the presence of air; uroporphyrinogens were only formed under anaerobic conditions, although substrate consumption was the same in the absence of oxygen as in its presence. 3. pH-dependence of both porphobilinogenase and deaminase was the same and a sharp optimum at pH 7.2 was obtained. Isomerase was heat-labile, but the presence of ammonium ions or porphobilinogen afforded some protection against inactivation. The action of several compounds added to the system was studied. Cysteine, thioglycollate, ammonium ions and hydroxylamine inhibited porphobilinogenase; certain concentrations of sodium and magnesium salts enhanced activity; some dicarboxylic acids and 2-methoxy-5-nitrotropone inhibited the deaminase. 4. delta-Aminolaevulate and ethionine in the culture media stimulated porphyrin synthesis and increased porphobilinogenase activity, whereas iron deficiency resulted in porphyrin accumulation. 5. The development of chlorophyll and porphobilinogenase on illumination of dark-grown callus was followed. 6. A hypothetical scheme is suggested for the enzymic synthesis of uroporphyrinogens from porphobilinogen.
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PMID:Studies on the porphobilinogen deaminase-uroporphyrinogen cosynthetase system of cultured soya-bean cells. 516 54

Enzyme preparations obtained from the mycelium of Aspergillus species broke down methionine by co-dissimilation. The deaminase and demethiolase activities of crude extracts were increased 100-fold by precipitation with (NH(4))(2)SO(4) and column chromatography on diethylaminoethyl cellulose. The enzyme acted on d-methionine but not on l-methionine. The enzyme was labile: it was inactivated by oxygen and ascorbic acid but ethylenediaminetetraacetic acid and mercaptoethanol preserved its activity. Enzyme activity decreased even at 4 and -30 C and was lost rapidly above 45 C. It was most rapid at 35 C and at pH 8.0 to 9.0. For the following reasons, it was concluded that deamination and demethiolation of methionine were effected by the same enzyme: both activities increased equally at each stage of purification; ammonia, methanethiol, and alpha-keto butyric acid were formed in amounts equivalent to the amount of methionine dissimilated; the K(m) and optimal pH for formation of both keto acid and methanethiol were the same; both activities remained in the same fractions that were separated by electrophoresis and the activities were equivalent. The purified enzyme demethiolated alpha-keto methionine and alpha-hydroxy methionine and split the sulfur linkage of ethionine but did not cleave cystathionine. Few amino acids were deaminated. The enzyme was sensitive to some carbonyl and sulfhydryl reagents and was relatively insensitive to heavy metals other than Hg(++). The K(m) was 1.3 x 10(-3) to 1.5 x 10(-3)m at pH 7.0. No requirement for cofactors was noted, and attempts to dissociate the enzyme, including dialysis with hydroxylamine, were unsuccessful.
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PMID:Dissimilation of methionine by a demethiolase of Aspergillus species. 537 Feb 77

The heat-stable polypeptide ATP-dependent proteolysis factor 1 (APF-1) of the reticulocyte proteolytic system forms covalent compounds with proteins in an ATP-requiring reaction. APF-1 and lysozyme, a good substrate for ATP-dependent proteolysis, form multiple conjugates, as was shown by comigration of label from each upon gel electrophoresis. Multiple bands were also seen with other substrates of the ATP-dependent proteolytic system, such as globin or alpha-lactalbumin. Analysis of the ratio of APF-1 to lysozyme radioactivities and of the molecular weights of the bands indicated that they consist of increasing numbers of the APF-1 polypeptide bound to one molecule of lysozyme. The covalent linkage is probably of an isopeptide nature, because it is stable to hydroxylamine and alkali, and polylysine is able to give conjugates of APF-1. Removal of ATP after formation of the 125I-labeled APF-1 conjugates with endogenous proteins caused the regeneration of APF-1, indicating presence of an amidase. This reaction is thought to compete with proteases that may act on APF-1-protein conjugates, especially those containing several APF-1 ligands. A sequence of reactions in which the linkage of APF-1 to the substrate is followed by the proteolytic breakdown of the substrate is proposed to explain the role of ATP.
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PMID:Proposed role of ATP in protein breakdown: conjugation of protein with multiple chains of the polypeptide of ATP-dependent proteolysis. 699 Apr 14


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