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)

PcsB is a protein of unknown function(s) that influences the cell morphology of several pathogenic species of streptococcus. PcsB contains a CHAP (cysteine, histidine-dependent amidohydrolase/peptidase) domain found in bacterial murein hydrolases; however, direct links between steps in cell wall biosynthesis and PcsB function(s) have not been demonstrated. We show here that pcsB is essential in the human respiratory pathogen, Streptococcus pneumoniae, that depletion of PcsB is bacteriostatic and that alanine substitutions in the conserved cysteine and histidine residues of the CHAP domain appear to be lethal. We stained wild-type parent and mutant bacteria deficient in expression of PcsB with fluorescent vancomycin and DAPI to determine patterns of cell wall synthesis and nucleoid segregation respectively. The wild-type parent strain exhibited ordered, simultaneous septal and equatorial cell wall synthesis. In contrast, reduced expression of PcsB resulted in formation of long chains of cells in which peptidoglycan synthesis occurred at nearly every division septum and cell equator. Severe depletion of PcsB led to abnormal, uncontrolled cell wall synthesis at misplaced septa and around large cells. Together, these physiological properties are consistent with a role for PcsB as a murein hydrolase that balances the extent of cell wall synthesis in S. pneumoniae. Finally, we show that the defects in morphology and cell wall synthesis that result from depletion of PcsB strongly resemble those caused by depletion of the essential VicRK two component regulatory system (TCS). This result and the essentiality of pcsB support the hypothesis that the essentiality of the VicRK TCS results from its positive regulation of PcsB expression.
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PMID:Defective cell wall synthesis in Streptococcus pneumoniae R6 depleted for the essential PcsB putative murein hydrolase or the VicR (YycF) response regulator. 1530 19

Biochemical characterization of the purified bile salt hydrolase (BSH) from Bifidobacterium bifidum ATCC 11863 revealed some distinct characteristics not observed in other species of Bifidobacterium. The bsh gene was cloned from B. bifidum, and the DNA flanking the bsh gene was sequenced. Comparison of the deduced amino acid sequence of the cloned gene with previously known sequences revealed high homology with BSH enzymes from several microorganisms and penicillin V amidase (PVA) of Bacillus sphaericus. The proposed active sites of PVA were highly conserved, including that of the Cys-1 residue. The importance of the SH group in the N-terminal cysteine was confirmed by substitution of Cys with chemically and structurally similar residues, Ser or Thr, both of which resulted in an inactive enzyme. The transcriptional start point of the bsh gene has been determined by primer extension analysis. Unlike Bifidobacterium longum bsh, B. bifidum bsh was transcribed as a monocistronic unit, which was confirmed by Northern blot analysis. PCR amplification with the type-specific primer set revealed the high level of sequence homology in their bsh genes within the species of B. bifidum.
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PMID:Cloning and characterization of the bile salt hydrolase genes (bsh) from Bifidobacterium bifidum strains. 1534 49

Acylase 1 from rat kidney catalyzes the hydrolysis of acyl-amino acids. Sequence alignment has shown that this enzyme belongs to the metalloprotein family M20. Site-directed mutagenesis experiments led to the identification of one functionally important amino acid residue located near one of the zinc coordinating residues, which play a critical role in the enzymatic activity. The D82N- and D82E-substituted forms showed no significant activity and very low activity, respectively, along with a loss of zinc coordination. Molecular modelling investigations indicated a putative role of D82 in ensuring a proper protonation of catalytic histidine. In addition, none of the five cysteine residues present in the rat kidney acylase 1 sequence seemed involved in the catalytic process: the loss of activity induced by the C294A substitution was probably due to a conformational change in the 3D structure.
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PMID:Site-directed mutagenesis and molecular modelling studies show the role of Asp82 and cysteines in rat acylase 1, a member of the M20 family. 1579 16

The controlled and partial modification of epoxy groups of Eupergit C and EP-Sepabeads with sodium sulfide has permitted the preparation of thiol-epoxy supports. Their use allowed not only the specific immobilization of enzymes through their thiol groups via thiol-disulfide interchange, but also enzyme stabilization via multipoint covalent attachment. Penicillin G acylase (PGA) from Escherichia coli and lipase from Rhizomucor miehei were used as model enzymes. Both enzymes lacked exposed cysteine residues, but were introduced via chemical modification under very mild conditions. In the first moments of the immobilization, a certain percentage of immobilized protein could be released from the support by incubation with DTT; this confirms that the first step was via a thiol-disulfide interchange. Moreover, the promotion of some further epoxy-enzyme bonds was confirmed because no enzyme release was detected after some immobilization time by incubation with DTT. In the case of the heterodimeric PGA, it was possible to demonstrate the formation of at least one epoxy bond per enzyme subunit by analyzing with SDS-PAGE the supernatants obtained after boiling the enzyme derivatives in the presence of mercaptoethanol and SDS. Thermal inactivation studies showed that these multipoint enzyme-support attachments promoted an increase in the stability of the immobilized enzymes. In both cases, the stabilization factor was around 12-15-fold comparing optimal derivatives with their just-thiol immobilized counterparts.
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PMID:Stabilization of enzymes by multipoint immobilization of thiolated proteins on new epoxy-thiol supports. 1581 62

Bacterial bile salt hydrolases catalyze the degradation of conjugated bile acids in the mammalian gut. The crystal structures of conjugated bile acid hydrolase (CBAH) from Clostridium perfringens as apoenzyme and in complex with taurodeoxycholate that was hydrolyzed to the reaction products taurine and deoxycholate are described here at 2.1 and 1.7 A resolution, respectively. The crystal structures reveal close relationship between CBAH and penicillin V acylase from Bacillus sphaericus. This similarity together with the N-terminal cysteine classifies CBAH as a member of the N-terminal nucleophile (Ntn) hydrolase superfamily. Both crystal structures show an identical homotetrameric organization with dihedral (D(2) or 222) point group symmetry. The structure analysis of C. perfringens CBAH identifies critical residues in catalysis, substrate recognition, and tetramer formation which may serve in further biochemical characterization of bile acid hydrolases.
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PMID:Conjugated bile acid hydrolase is a tetrameric N-terminal thiol hydrolase with specific recognition of its cholyl but not of its tauryl product. 1582 32

A bacterium which degrades urethane compounds was isolated and identified as Rhodococcus equi strain TB-60. Strain TB-60 degraded toluene-2,4-dicarbamic acid dibutyl ester (TDCB) and accumulated toluene diamine as the degradation product. The enzyme which cleaves urethane bond in TDCB was strongly induced by acetanilide. The purified enzyme (urethane hydrolase) was found to be homogeneous on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The molecular weight was estimated to be 55 kDa. The optimal temperature and pH were 45 degrees C and 5.5, respectively. The enzyme hydrolyzed aliphatic urethane compound as well as aromatic ones. The activity was inhibited by HgCl(2), p-chrolomercuribenzoic acid, and phenylmethylsulfonyl fluoride, suggesting that cysteine and/or serine residues play an important role in the activity. The enzyme catalyzed the hydrolysis of anilides, amides, and esters as well as TDCB. It was characterized as a novel amidase/esterase, differing in some properties from other known amidases/esterases.
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PMID:Isolation of a bacterium that degrades urethane compounds and characterization of its urethane hydrolase. 1604 75

We have determined the crystal structures of three homologous proteins from the pathogenic protozoans Leishmania donovani, Leishmania major, and Trypanosoma cruzi. We propose that these proteins represent a new subfamily within the isochorismatase superfamily (CDD classification cd004310). Their overall fold and key active site residues are structurally homologous both to the biochemically well-characterized N-carbamoylsarcosine-amidohydrolase, a cysteine hydrolase, and to the phenazine biosynthesis protein PHZD (isochorismase), an aspartyl hydrolase. All three proteins are annotated as mitochondrial-associated ribonuclease Mar1, based on a previous characterization of the homologous protein from L. tarentolae. This would constitute a new enzymatic activity for this structural superfamily, but this is not strongly supported by the observed structures. In these protozoan proteins, the extended active site is formed by inter-subunit association within a tetramer, which implies a distinct evolutionary history and substrate specificity from the previously characterized members of the isochorismatase superfamily. The characterization of the active site is supported crystallographically by the presence of an unidentified ligand bound at the active site cysteine of the T. cruzi structure.
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PMID:Crystal structures and proposed structural/functional classification of three protozoan proteins from the isochorismatase superfamily. 1619 69

We identified an aldoxime dehydratase (Oxd) gene in the 5'-flanking region of the nitrile hydratase-amidase gene cluster in the photoreactive iron-type nitrile hydratase-producer, Rhodococcus sp. N-771. The enzyme showed 96.3%, 77.6%, and 30.4% identities with the Oxds of Rhodococcus globerulus A-4, Pseudomonas chlororaphis B23, and Bacillus sp. OxB-1, respectively. The enzyme was expressed in Escherichia coli under the control of the lac- or T7 promoters in its intact and His6-tagged forms, purified, and characterized. The enzyme had heme b as a prosthetic group, catalyzed a stoichiometric dehydration of aldoxime into nitrile, and exhibited the highest activity at neutral pH and at around 30 degrees C similar to the known Oxd from Bacillus sp. OxB-1. The activity was enhanced by reducing agents, such as Na2S, Na2S2(O4), 2-mercaptoethanol, and L-cysteine and supplementary additions of electron acceptors such as flavins, sulfite ion, and vitamin K3. The effect of various chemicals on the enzyme activity was different in the presence and absence of the reducing reagent, Na2S. The enzyme preferentially acts on aliphatic-type substrates and the substrate specificity of the enzyme coincides with that reported for nitrile hydratase produced by the strain.
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PMID:Aldoxime dehydratase co-existing with nitrile hydratase and amidase in the iron-type nitrile hydratase-producer Rhodococcus sp. N-771. 1623 24

From a forward genetic screen for phagocytosis mutants in Drosophila melanogaster, we identified a mutation that affects peptidoglycan recognition protein (PGRP) SC1a and impairs the ability to phagocytose the bacteria Staphylococcus aureus, but not Escherichia coli and Bacillus subtilis. Because of the differences in peptidoglycan peptide linkages in these bacteria, our data suggest that PGRP-SC1a is necessary for recognition of the Lys-type peptidoglycan typical of most Gram(+) bacteria. PGRP-SC1a mutants also fail to activate the Toll/NF-kappaB signaling pathway and are compromised for survival after S. aureus infection. This mutant phenotype is the first found for an N-acetylmuramoyl-l-alanine amidase PGRP that cleaves peptidoglycan at the lactylamide bond between the glycan backbone and the crosslinking stem peptides. By generating transgenic rescue flies that express either wild-type or a noncatalytic cysteine-serine mutant PGRP-SC1a, we find that PGRP-SC1a amidase activity is not necessary for Toll signaling, but is essential for uptake of S. aureus into the host phagocytes and for survival after S. aureus infection. Furthermore, we find that the PGRP-SC1a amidase activity can be substituted by exogenous addition of free peptidoglycan, suggesting that the presence of peptidoglycan cleavage products is more important than the generation of cleaved peptidoglycan on the bacterial surface for PGRP-SC1a mediated phagocytosis.
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PMID:The peptidoglycan recognition protein PGRP-SC1a is essential for Toll signaling and phagocytosis of Staphylococcus aureus in Drosophila. 1640 37

APOBEC3G (APO3G) is a cytidine deaminase that restricts replication of vif-defective human immunodeficiency virus type 1 (HIV-1). Like other members of the cellular deaminase family, APO3G has the propensity to form homo-multimers. In the current study, we investigated the functional determinants for multimerization of human APO3G and studied the role of APO3G multimerization for catalytic activity, virus encapsidation, and antiviral activity. We found that human APO3G is capable of forming multimeric complexes in transfected HeLa cells. Interestingly, multimerization of APO3G was exquisitely sensitive to RNase treatment, suggesting that interaction of APO3G subunits is facilitated or stabilized by an RNA bridge. Mutation of a conserved cysteine residue (C97) that is part of an N-terminal zinc-finger motif in APO3G abolished multimerization of APO3G; however, the C97 mutation inhibited neither in vitro deaminase activity nor antiviral function of APO3G. These results suggest that monomeric APO3G is both catalytically active and has antiviral activity. Interference studies employing either catalytically inactive or packaging-incompetent APO3G variants suggest that wild-type APO3G is packaged into HIV-1 particles in monomeric form. These results provide novel insights into the catalytic function and antiviral property of APO3G and demonstrate an important role for C97 in the RNA-dependent multimerization of this protein.
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PMID:Monomeric APOBEC3G is catalytically active and has antiviral activity. 1664 Dec 60


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