Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.5.1.4 (deaminase)
 5,113 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The ability of neurons to metabolize sulfur-containing compounds to cysteine was investigated using as indicator the glutathione content in neuron-rich primary cultures derived from the brains of embryonal rats. The-glutathione content of these cultures was doubled during a 4-h incubation in a minimal medium containing cysteine, glutamine and glycine. In contrast, absence of cysteine or replacement of cysteine by methionine or 2-oxothiazolidine-4-carboxylate failed to increase the glutathione content of cultured neurons. Besides cysteine, N-acetylcysteine (NAC) also caused in the millimolar range, a concentration-dependent increase in the neuronal glutathione content during a 4-h incubation. These data suggest that neurons in culture, contain an acylase activity which allows them to generate from extracellular NAC as precursor intracellular cysteine in concentrations sufficient for glutathione synthesis. In contrast, generation of cysteine from 2-oxothiazolidine-4-carboxylate by the reaction of 5-oxoprolinase or from methionine by the transsulfuration pathway appears not to take place in these cultured neurons.
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PMID:N-acetylcysteine, but not methionine or 2-oxothiazolidine-4-carboxylate, serves as cysteine donor for the synthesis of glutathione in cultured neurons derived from embryonal rat brain. 1002 62

The nitrile hydratase (NHase) from Rhodococcus sp. N-771 is a photoreactive enzyme that is inactivated on nitrosylation of the non-heme iron center and activated on photo-dissociation of nitric oxide (NO). The nitrile hydratase operon consists of six genes encoding NHase regulator 2, NHase regulator 1, amidase, NHase alpha subunit, NHase beta subunit and NHase activator. We overproduced the NHase in Escherichia coli using a T7 expression system. The NHase was functionally expressed in E. coli only when the NHase activator encoded downstream of the beta subunit gene was co-expressed and the transformant was grown at 30 degrees C or less. A ligand cysteine, alphaCys112, of the recombinant NHase was also post-translationally modified to a cysteine-sulfinic acid similar to for the native NHase. Although another modification of alphaCys114 could not be identified because of the instability under acidic conditions, the recombinant NHase could be reversibly inactivated by nitric oxide.
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PMID:Functional expression of nitrile hydratase in Escherichia coli: requirement of a nitrile hydratase activator and post-translational modification of a ligand cysteine. 1010 Dec 82

The dried latex of the mountain papaya, Carica candamarcensis, was chromatographed on CM-Sephadex C50, giving rise to three peaks (CCI, CCII and CCIII) with amidase activity on N-alpha-benzoyl-DL-arginine-4-nitroanilide. The less basic, most active, peak, CCI, was separated into two components, CCIa and CCIb, by reverse-phase HPLC under denaturing conditions. The primary structures of CCIa and CCIb are presented. They were deduced from sequence analysis of the whole proteins and peptides resulting from enzymatic digestions. Both proteinases are made of 213 amino acid residues, CCIb sharing 88-89% similarity with the three subvariants (G90/R212, E90/R212, E90/K212) of CCIa. 139-140 amino acid residues (65.8%) of CCIa and 141 residues (66.5%) of CCIb are common to papain. The seven cysteine residues are aligned with those of papain and the catalytic triad (Cys25, His159, Asn175) of all cysteine peptidases of the papain family is conserved. The similarity with the other cysteine proteases from Carica papaya is discussed.
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PMID:Isolation and primary structure of the CCI papain-like cysteine proteinases from the latex of Carica candamarcensis hook. 1035 34

Mercapturates (S-substituted N-acetyl-L-cysteines) are terminal metabolites formed by the glutathione-dependent metabolism of electrophilic xenobiotics, including haloalkenes. Acylases catalyze the hydrolysis of N-acyl-L-amino acids, including many xenobiotic-derived mercapturates, to give fatty acids and amino acids as products. Although several acylases have been identified, the acylases that catalyze the deacetylation of the haloalkene-derived mercapturates have not been identified and characterized. Acylase I catalyzes the deacetylation of some haloalkene-derived mercapturates, including S-(1,1,2, 2-tetrafluoroethyl)-N-acetyl-L-cysteine, S-(2-chloro-1,1, 2-trifluoroethyl)-N-acetyl-L-cysteine, and S-(2-bromo-1,1, 2-trifluoroethyl)-N-acetyl-L-cysteine [Uttamsingh, V., et al. (1998) Chem. Res. Toxicol. 11, 800-809]. In the studies presented here, we identified a rat kidney acylase that catalyzed the hydrolysis of the haloalkene-derived mercapturates S-(1, 2-dichlorovinyl)-N-acetyl-L-cysteine, S-(1,2,3,4,4-pentachloro-1, 3-butadienyl)-N-acetyl-L-cysteine, and S-(2,2-dibromo-1, 1-difluoroethyl)-N-acetyl-L-cysteine. The substrate selectivity and amino acid sequence of the purified rat kidney acylase were studied. Although the sequence of the purified rat kidney acylase was somewhat identical with that of aspartoacylase, it did not catalyze the hydrolysis of N-acetyl-L-aspartate.
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PMID:Acylase-catalyzed deacetylation of haloalkene-derived mercapturates. 1052 69

Acylases catalyze the hydrolysis of a range of S-substituted N-acetyl-L-cysteines. The hydrolysis of N-acetyl-L-cysteine is catalyzed by cytosolic acylase I, and activity is present in human endothelial cells and rat lung, intestinal, and liver homogenates. Many haloalkenes are metabolized to mercapturates, which also undergo acylase-catalyzed hydrolysis. The acylases that catalyze the deacetylation of N-acetyl-L-cysteine and several haloalkene-derived mercapturates have been recently identified: acylase I catalyzes the deacetylation of N-acetyl-L-cysteine and some haloalkene-derived mercapturates whereas an acylase purified from rat kidney cytosol catalyzes the deacetylation of a distinct set of substrates, including several haloalkene-derived mercapturates. The objective of these studies was to examine the tissue and subcellular localization of acylase I and purified rat kidney acylase. Immunoblotting showed the presence of immunoreactive acylase I and purified rat kidney acylase in rat kidney, liver, lung, and brain. Both acylases were identified by immunohistochemistry in several rat organs, including kidney, liver, lung, brain, stomach, intestines, adrenals, pancreas, and testis, indicating that acylase activity is widespread in rat tissues.
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PMID:Immunohistochemical localization of the acylases that catalyze the deacetylation of N-acetyl-L-cysteine and haloalkene-derived mercapturates. 1082 Jan 33

Methanethiosulfonate reagents may be used to introduce virtually unlimited structural modifications in enzymes via reaction with the thiol group of cysteine. The covalent coupling of enantiomerically pure (R) and (S) chiral auxiliary methanethiosulfonate ligands to cysteine mutants of subtilisin Bacillus lentus induces spectacular changes in catalytic activity between diastereomeric enzymes. Amidase and esterase kinetic assays using a low substrate approximation were used to establish kcat/KM values for the chemically modified mutants, and up to 3-fold differences in activity were found between diastereomeric enzymes. Changing the length of the carbon chain linking the phenyl or benzyl oxazolidinone ligand to the mutant N62C by a methylene unit reverses which diastereomeric enzyme is more active. Similarly, changing from a phenyl to benzyl oxazolidinone ligand at S166C reverses which diastereomeric enzyme is more active. Chiral modifications at S166C and L217C give CMMs having both high esterase kcat/KM's and high esterase to amidase ratios with large differences between diastereomeric enzymes.
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PMID:Covalent modification of subtilisin Bacillus lentus cysteine mutants with enantiomerically pure chiral auxiliaries causes remarkable changes in activity. 1100 41

Mycothiol is a novel thiol produced only by actinomycetes and is the major low-molecular-weight thiol in mycobacteria. Mycothiol was previously shown to be synthesized from 1-D-myo-inosityl-2-amino-2-deoxy-alpha-D-glucopyranoside by ligation with cysteine followed by acetylation. A novel mycothiol-dependent detoxification enzyme, mycothiol conjugate amidase, was recently identified in Mycobacterium smegmatis and shown to have a homolog, Rv1082, in Mycobacterium tuberculosis. In the present study we found that a protein encoded by the M. tuberculosis open reading frame Rv1170, a homolog of Rv1082, possesses weak mycothiol conjugate amidase activity but shows substantial deacetylation activity with 1-D-myo-inosityl-2-acetamido-2-deoxy-alpha-D-glucopyranoside (GlcNAc-Ins), a hypothetical mycothiol biosynthetic precursor. The availability of this protein enabled us to develop an assay for GlcNAc-Ins, which was used to demonstrate that GlcNAc-Ins is present in M. smegmatis at a level about twice that of mycothiol. It was shown that GlcNAc-Ins is absent in mycothiol-deficient mutant strain 49 of M. smegmatis and that this strain can concentrate GlcNAc-Ins from the medium and convert it to mycothiol. This demonstrates that GlcNAc-Ins is a key intermediate in the pathway of mycothiol biosynthesis. Assignment of Rv1170 as the gene coding the deacetylase in the M. tuberculosis genome represents the first identification of a gene of the mycothiol biosynthesis pathway. The presence of a large cellular pool of substrate for this enzyme suggests that it may be important in regulating mycothiol biosynthesis.
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PMID:N-Acetyl-1-D-myo-inosityl-2-amino-2-deoxy-alpha-D-glucopyranoside deacetylase (MshB) is a key enzyme in mycothiol biosynthesis. 1109 56

In an attempt to investigate the molecular basis of pyrazinamide hydrolysis by the PncA protein from Mycobacterium tuberculosis, we determined the pyrazinamidase activity of nine PncA mutants bearing a single amino acid substitution. Among them, three mutants (D8G, K96T and S104R) had virtually no activity (< or =0.004 unit/mg), five (F13S, T61P, P69L, Y103S and A146V) retained a low level of activity (0.06-0.25 unit/mg) and one (T167L) exhibited a wild-type activity (1.51 units/mg). The possible structural effects of these substitutions were assessed by analysing a three-dimensional model of the PncA protein constructed on the basis of the crystal structure of the N-carbamoylsarcosine amidohydrolase (CSHase) from Arthrobacter sp., an amidohydrolase which was found by hydrophobic cluster analysis to be closely related to PncA. In the PncA model, five of the mutated residues, Asp-8, Phe-13, Lys-96, Tyr-103 and Ser-104, were located within a 6 A sphere around the cysteine residue Cys-138, which could be the counterpart of the active cysteine residue Cys-177 found in the CSHase. Among the remaining mutated residues, Thr-61, Pro-69 and Ala-146 were found to be more distant from Cys-138 but were associated with structural elements contributing to the catalytic centre, whereas Thr-167 was situated in an alpha-helix located far from the putative active site. These data suggest that the decrease in pyrazinamidase activity observed in the PncA mutant proteins is well correlated with the structural modifications the mutations can cause in the environment of the putative active cysteine Cys-138.
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PMID:Study of the structure-activity relationships for the pyrazinamidase (PncA) from Mycobacterium tuberculosis. 1117 Oct 40

Species differences in the biotransformation of N-acetyl-L-cysteine (NAC) have been investigated to evaluate the usefulness of NAC as a constituent in parenteral nutrition solutions in place of cysteine. The activity of NAC-deacetylating enzyme (acylase) was measured in various tissues of different species (rat, rabbit, dog, monkey, and man). Acylase activity was highest in the kidney in all species studied. Enzyme activity in the liver was 10 %-22 % of that in the kidney in the rat, rabbit, monkey, and man, but almost no hepatic activity was seen in the dog. NAC-deacetylating activity was very low in other organs. The tissue distribution of acylase I was determined by Western blotting and an immunohistochemical method employing specific antibody against porcine acylase I (EC 3.5.1.14). The immunoblotting study showed a 46-kDa protein band corresponding to porcine acylase I in the kidney of all species. In liver cytosol, 46 kDa and/or 29 kDa bands were observed in the rat, rabbit, monkey, and man, but not in the dog. In the immunohistochemical study, positive staining with anti-acylase I antibody was observed clearly in the renal proximal tubules in the monkey and man. These results suggested that the kidney and liver were the main organs responsible for the biotransformation of NAC to cysteine in mammals other than the dog.
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PMID:Tissue distribution of and species differences in deacetylation of N-acetyl-L-cysteine and immunohistochemical localization of acylase I in the primate kidney. 1184 84

DL-2-amino-Delta(2)-thiazolin-4-carbonic acid (DL-ATC) is a substrate for cysteine synthesis in some bacteria, and this bioconversion has been utilized for cysteine production in industry. We cloned a DNA fragment containing the genes involved in the conversion of L-ATC to L-cysteine from Pseudomonas sp. strain BS. The introduction of this DNA fragment into Escherichia coli cells enabled them to convert L-ATC to cysteine via N-carbamyl-L-cysteine (L-NCC) as an intermediate. The smallest recombinant plasmid, designated pTK10, contained a 2.6-kb insert DNA fragment that has L-cysteine synthetic activity. The nucleotide sequence of the insert DNA revealed that two open reading frames (ORFs) encoding proteins with molecular masses of 19.5 and 44.7 kDa were involved in the L-cysteine synthesis from DL-ATC. These ORFs were designated atcB and atcC, respectively, and their gene products were identified by overproduction of proteins encoded in each ORF and by the maxicell method. The functions of these gene products were examined using extracts of E. coli cells carrying deletion derivatives of pTK10. The results indicate that atcB and atcC are involved in the conversion of L-ATC to L-NCC and the conversion of L-NCC to cysteine, respectively. atcB was first identified as a gene encoding an enzyme that catalyzes thiazolin ring opening. AtcC is highly homologous with L-N-carbamoylases. Since both enzymes can only catalyze the L-specific conversion from L-ATC to L-NCC or L-NCC to L-cysteine, it is thought that atcB and atcC encode L-ATC hydrolase and N-carbamyl-L-cysteine amidohydrolase, respectively.
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PMID:Genes from Pseudomonas sp. strain BS involved in the conversion of L-2-amino-Delta(2)-thiazolin-4-carbonic acid to L-cysteine. 1197 87


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