Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Intramolecular Transferases”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 235 records · Page 13Linked to original sources
PubMed2004

Synthesis of a novel class of sulfonium ions as potential inhibitors of UDP-galactopyranose mutase.

Two sulfonium salts of 1,4-anhydro-4-thio-D-galactitol, with structures related to the known sulfonium salt glycosidase inhibitor, salacinol, have been synthesized as potential inhibitors of UDP-galactopyranose mutase. The synthetic strategy relies on the alkylation reaction of 1,4-anhydro-2,3,5,6-tetra-O-benzyl-4-thio-D-galactitol at the sulfur atom with 2,4-O-benzylidene-D- or -L-erythritol-1,3-cyclic sulfate. In each case, the reaction proceeded stereoselectively to yield only one stereoisomer at the stereogenic sulfur atom. The effect of the polar solvent, 1,1,1,3,3,3-hexafluoroisopropanol (HFIP), in promoting high-yielding reactions is highlighted. The target compounds are then obtained by hydrogenolysis.

Enzyme Inhibitors↗
PubMed2004

Synthesis of novel ammonium and selenonium ions and their evaluation as inhibitors of UDP-galactopyranose mutase.

The syntheses of two ammonium salts of 1,4-dideoxy-1,4-imino-d-galactitol containing erythritol sulfate side chains are described. The parent compound is a known inhibitor of the enzyme UDP-galactopyranose mutase (UDP-galactopyranose furanomutase, E.C. 5.4.99.9), which is responsible for the conversion of UDP-galactopyranose into UDP-galactofuranose and is presumably protonated in its active form. The side chain was chosen because it is present in a known sulfonium ion alpha-glucosidase inhibitor, salacinol. The syntheses of the selenonium analogues derived from 1,4-dideoxy-1,4-seleno-d-galactitol are also described. The synthetic strategy in the syntheses of all four salts involved the nucleophilic attack of a protected derivative of the alditol at the least hindered carbon of 2,4-O-benzylidene d- or l-erythritol-1,3-cyclic sulfate to give adducts that were subsequently deprotected. The importance of different protecting groups used in the various syntheses is also highlighted. Enzyme inhibition assays carried out on these compounds, and the corresponding sulfonium ions synthesized previously, show that they are poor inhibitors of UDP-galactopyranose mutase.

Carbohydrate Conformation↗
PubMed2004

Cloning, sequencing, and functional analysis of the biosynthetic gene cluster of macrolactam antibiotic vicenistatin in Streptomyces halstedii.

Vicenistatin, an antitumor antibiotic isolated from Streptomyces halstedii, is a unique 20-membered macrocyclic lactam with a novel aminosugar vicenisamine. The vicenistatin biosynthetic gene cluster (vin) spanning approximately 64 kbp was cloned and sequenced. The cluster contains putative genes for the aglycon biosynthesis including four modular polyketide synthases (PKSs), glutamate mutase, acyl CoA-ligase, and AMP-ligase. Also found in the cluster are genes of NDP-hexose 4,6-dehydratase and aminotransferase for vicenisamine biosynthesis. For the functional confirmation of the cluster, a putative glycosyltransferase gene product, VinC, was heterologously expressed, and the vicenisamine transfer reaction to the aglycon was chemically proved. A unique feature of the vicenistatin PKS is that the loading module contains only an acyl carrier protein domain, in contrast to other known PKS-loading modules containing certain activation domains. Activation of the starter acyl group by separate polypeptides is postulated as well.

Aminoglycosides↗
PubMed2004

Conversion of squalene to the pentacarbocyclic hopene.

The membrane protein squalene-hopene cyclase was cocrystallized with 2-azasqualene and analyzed by X-ray diffraction to 2.13 A resolution. The conformation of this close analog was clearly established, and it agreed with the common textbook presentation. The bound squalene undergoes only small conformational changes during the formation of rings A through D, thus requiring no intermediate. However, ring E formation is hindered by an entropic barrier, which may explain its absence in the steroids. The structure analysis revealed a mobile region between the active center cavity and the membrane, which may melt, opening a passage for squalene and hopene.

Bacillaceae↗
PubMed2004

Profound insights into squalene cyclization.

In this issue of Chemistry & Biology, our understanding of the formation of pentacyclic hopene from the linear squalene is enhanced by an X-ray structure of a complex between squalene-hopene cyclase and the substrate analog 2-azasqualene.

Crystallography, X-Ray↗
PubMed2006

Chemical probes of UDP-galactopyranose mutase.

Many pathogenic prokaryotes and eukaryotes possess the machinery required to assemble galactofuranose (Galf)-containing glycoconjugates; these glycoconjugates can be critical for virulence or viability. Accordingly, compounds that block Galf incorporation may serve as therapeutic leads or as probes of the function of Galf-containing glycoconjugates. The enzyme UDP-galactopyranose mutase (UGM) is the only known generator of UDP-galactofuranose, the precursor to Galf residues. We previously employed a high-throughput fluorescence polarization assay to investigate the Klebsiella pneumoniae UGM. We demonstrate the generality of this assay by extending it to UGM from Mycobacterium tuberculosis. To identify factors influencing binding, we synthesized a directed library containing a 5-arylidene-2-thioxo-4-thiazolidinone core, a structure possessing features common to ligands for both homologs. Our studies offer a blueprint for identifying inhibitors of the growing family of UGM homologs and provide insight into UGM inhibition.

Binding Sites↗
PubMed2004

Umbelliferone aminoalkyl derivatives, a new class of squalene-hopene cyclase inhibitors.

The synthesis is described of several aminoalkyl derivatives of coumarin, obtained in good yields under microwave or high-intensity ultrasound irradiation. These compounds proved uniformly active as inhibitors of squalene-hopene cyclase (SHC) from Alicyclobacillus acidocaldarius. Their design stemmed from our recent finding that the umbelliferone nucleus acquires inhibitory properties towards SHC when functionalized with a suitable chain such as the omega-epoxyfarnesyl group. Under our experimental conditions the most active ones, such as 7-(4'-allylmethylamino-but-2-ynyloxy)chromen-2-one (IC(50) 0.75 mM), approached the potency of anticholesteremic drug Ro 48-8071 (IC(50) 0.35 mM), an effective inhibitor of both squalene- and oxidosqualene-cyclases (OSC). Tests are in progress to determine their efficacy on different eukaryotic OSCs.

Animals↗
PubMed2005

Histopathologic findings after treatment with different oxidosqualene cyclase (OSC) inhibitors in hamsters and dogs.

In order to investigate the toxicity of oxidosqualene cyclase (OSC) inhibitors, hamsters and dogs were treated up to 13 weeks with three different agents of this compound class. Subacute treatment (< or = 4 weeks) of hamsters and dogs with OSC inhibitors produced a similar spectrum of histopathologic lesions, which have previously been described for compounds of this pharmacological class. In the hamster, other lesions were produced only after subchronic treatment (13 weeks). After subacute treatment, histopathologic findings were observed in the eye, skin and forestomach. Lesions in the eye were characterized as proliferation, swelling and degeneration of lens fiber cells. This finding was considered to be an early stage in cataract development. In the skin and forestomach, squamous cell hyperplasia and hyperkeratosis accompanied by a mixed inflammation was observed. After subchronic treatment of hamsters, histopathologic findings were observed in the eye, skin, forestomach, testes, epididymides, prostate gland, seminal vesicles and long bones. Lesions in the eye were characterized as cataracts. Squamous cell hyperplasia accompanied by a diffuse hyperkeratosis and a mixed inflammation was seen in the skin and forestomach epithelium. In addition, testes lesions were characterized as testicular atrophy, generalized germ cell depletion, germ cell degeneration and tubular collapse. Atrophy, oligospermia and lumenal germ cells/cell debris were found in the epididymides. The prostate gland and seminal vesicles were decreased in size (atrophy). The bone lesions were characterized as a failure of enchondral ossification causing variable widening of the growth plate and a failure to form primary bone trabecula (lesions resemble those found in rickets). To our knowledge, this is the first study describing the toxicity of OSC inhibitors after subchronic treatment in hamsters and dogs. As all adverse effects described in this report are considered to be due to an exaggeration of the desired biochemical mechanism of action at high dose levels, a decrease of the systemic exposure by the use of more hepatoselective OSC inhibitors is expected to reduce the probability of these adverse effects in humans.

Animals↗
PubMed2004

X-ray structure of tRNA pseudouridine synthase TruD reveals an inserted domain with a novel fold.

Pseudouridine synthases catalyse the isomerisation of uridine to pseudouridine in structural RNA. The pseudouridine synthase TruD, that modifies U13 in tRNA, belongs to a recently identified and large family of pseudouridine synthases present in all kingdoms of life. We report here the crystal structure of Escherichia coli TruD at 2.0 A resolution. The structure reveals an overall V-shaped molecule with an RNA-binding cleft formed between two domains: a catalytic domain and an insertion domain. The catalytic domain has a fold similar to that of the catalytic domains of previously characterised pseudouridine synthases, whereas the insertion domain displays a novel fold.

Amino Acid Sequence↗
PubMed2005

Characterisation of plant tocopherol cyclases and their overexpression in transgenic Brassica napus seeds.

Tocopherols, collectively known as vitamin E, are only synthesised in photosynthetic organisms. Tocopherol cyclase (TC) catalyses the formation of the chromanol headgroup of the various tocopherol isoforms. TCs from Arabidopsis and maize (Zea mays) were expressed in Escherichia coli and purified. Analysis of the enzymatic properties revealed similarities but also differences between the two enzymes. Overexpression of chimeric TC gene constructs in developing seeds of transgenic rapeseed plants enhanced and modified the relative abundance of individual tocochromanol species in the seed oil, indicating a regulatory function of the enzyme in prenyllipid metabolism.

Arabidopsis↗
PubMed2006

Cloning and characterization of the erg1 gene of Trichoderma harzianum: effect of the erg1 silencing on ergosterol biosynthesis and resistance to terbinafine.

Trichoderma species are commonly used as biocontrol agents of different plant-pathogenic fungi. Terpene compounds are involved in the biocontrol process due to their antifungal properties (e.g., ergokonins and viridins) but additionally their structural function in the cell membranes (ergosterol) is essential. We report here the characterization of the T. harzianum erg1 gene, encoding a squalene epoxidase, a key enzyme in the biosynthesis of triterpene derivatives such as ergosterol. In T. harzianum the partial silencing of the erg1 gene gave rise to transformants with a higher level of sensitivity to terbinafine, an antifungal compound that acts specifically over the squalene epoxidase activity. In addition, these silenced transformants produced lower levels of ergosterol than the wild type strain. Finally, the silencing of the erg1 gene resulted in an increase in the expression level of the erg7 gene that encodes the oxidosqualene lanosterol-cyclase, another enzyme of the terpene biosynthesis pathway.

Antifungal Agents↗
PubMed2006

Vitamin K1 accumulation in tobacco plants overexpressing bacterial genes involved in the biosynthesis of salicylic acid.

Phylloquinone (Vitamin K(1)) is an essential component of the photosynthetic electron transfer. As isochorismate is required for the biosynthesis of Vitamin K(1), isochorismate synthase (ICS) activity is expected to be present in all green plants. In bacteria salicylic acid (SA) is synthesized via a two step pathway involving ICS and isochorismate pyruvate lyase (IPL). The effect of the introduction in tobacco plants of the bacterial ICS and IPL genes on the endogenous isochorismate pathway was investigated. Transgenic tobacco plants in which IPL was targeted to the chloroplast suffered severe growth retardation and had low Vitamin K(1) content. Probably because isochorismate was channeled towards SA production, the plants were no longer able to produce normal levels of Vitamin K(1). Transgenic tobacco plants in which the bacterial ICS was present in the chloroplast showed higher Vitamin K(1) contents than wild type plants.

Bacterial Proteins↗
PubMed2004

Crystal structure of the RluD pseudouridine synthase catalytic module, an enzyme that modifies 23S rRNA and is essential for normal cell growth of Escherichia coli.

Pseudouridine (5-beta-D-ribofuranosyluracil, Psi) is the most commonly found modified base in RNA. Conversion of uridine to Psi is performed enzymatically in both prokaryotes and eukaryotes by pseudouridine synthases (EC 4.2.1.70). The Escherichia coli Psi-synthase RluD modifies uridine to Psi at positions 1911, 1915 and 1917 within 23S rRNA. RluD also possesses a second function related to proper assembly of the 50S ribosomal subunit that is independent of Psi-synthesis. Here, we report the crystal structure of the catalytic module of RluD (residues 68-326; DeltaRluD) refined at 1.8A to a final R-factor of 21.8% (R(free)=24.3%). DeltaRluD is a monomeric enzyme having an overall mixed alpha/beta fold. The DeltaRluD molecule consists of two subdomains, a catalytic subdomain and C-terminal subdomain with the RNA-binding cleft formed by loops extending from the catalytic sub-domain. The catalytic sub-domain of DeltaRluD has a similar fold as in TruA, TruB and RsuA, with the location of the RNA-binding cleft, active-site and conserved, catalytic Asp residue superposing in all four structures. Superposition of the crystal structure of TruB bound to a T-stem loop with RluD reveals that similar RNA-protein interactions for the flipped-out uridine base would exist in both structures, implying that base-flipping is necessary for catalysis. This observation also implies that the specificity determinants for site-specific RNA-binding and recognition likely reside in parts of RluD beyond the active site.

Catalytic Domain↗
PubMed2004

Conformational flexibility of the C terminus with implications for substrate binding and catalysis revealed in a new crystal form of deacetoxycephalosporin C synthase.

Deacetoxycephalosporin C synthase (DAOCS) from Streptomyces clavuligerus catalyses the oxidative ring expansion of the penicillin nucleus into the nucleus of cephalosporins. The reaction requires dioxygen and 2-oxoglutarate as co-substrates to create a reactive iron-oxygen intermediate from a ferrous iron in the active site. The active enzyme is monomeric in solution. The structure of DAOCS was determined earlier from merohedrally twinned crystals where the last four C-terminal residues (308-311) of one molecule penetrate the active site of a neighbouring molecule, creating a cyclic trimeric structure in the crystal. Shortening the polypeptide chain from the C terminus by more than four residues diminishes activity. Here, we describe a new crystal form of DAOCS in which trimer formation is broken and the C-terminal arm is free. These crystals show no signs of twinning, and were obtained from DAOCS labelled with an N-terminal His-tag. The modified DAOCS is catalytically active. The free C-terminal arm protrudes into the solvent, and the C-terminal domain (residues 268-299) is rotated by about 16 degrees towards the active site. The last 12 residues (300-311) are disordered. Structures for various enzyme-substrate and enzyme-product complexes in the new crystal form confirm overlapping binding sites for penicillin and 2-oxoglutarate. The results support the notion that 2-oxoglutarate and dioxygen need to react first to produce an oxidizing iron species, followed by reaction with the penicillin substrate. The position of the penicillin nucleus is topologically similar in the two crystal forms, but the penicillin side-chain in the new non-twinned crystals overlaps with the position of residues 304-306 of the C-terminal arm in the twinned crystals. An analysis of the interactions between the C-terminal region and residues in the active site indicates that DAOCS could also accept polypeptide chains as ligands, and these could bind near the iron.

Amino Acid Sequence↗
PubMed2005

Crystal structures of Mycobacteria tuberculosis and Klebsiella pneumoniae UDP-galactopyranose mutase in the oxidised state and Klebsiella pneumoniae UDP-galactopyranose mutase in the (active) reduced state.

Uridine diphosphogalactofuranose (UDP-Galf) is the precursor of the d-galactofuranose sugar found in bacterial and parasitic cell walls, including those of many pathogens. UDP-Galf is made from UDP-galactopyranose by the enzyme UDP-galactopyranose mutase. The enzyme requires the reduced FADH- co-factor for activity. The structure of the Mycobacterium tuberculosis mutase with FAD has been determined to 2.25 A. The structures of Klebsiella pneumoniae mutase with FAD and with FADH- bound have been determined to 2.2 A and 2.35 A resolution, respectively. This is the first report of the FADH(-)-containing structure. Two flavin-dependent mechanisms for the enzyme have been proposed, one, which involves a covalent adduct being formed at the flavin and the other based on electron transfer. Using our structural data, we have examined the two mechanisms. The electron transfer mechanism is consistent with the structural data, not surprisingly, since it makes fewer demands on the precise positioning of atoms. A model based on a covalent adduct FAD requires repositioning of the enzyme active site and would appear to require the isoalloxazine ring of FADH- to buckle in a particular way. However, the FADH- structure reveals that the isoalloxazine ring buckles in the opposite sense, this apparently requires the covalent adduct to trigger profound conformational changes in the protein or to buckle the FADH- opposite to that seen in the apo structure.

Amino Acid Sequence↗
PubMed2006

Evolutionary relationship between initial enzymes of tetrapyrrole biosynthesis.

Glutamate-1-semialdehyde 2,1-aminomutase (GSAM) is the second enzyme in the C(5) pathway of tetrapyrrole biosynthesis found in most bacteria, in archaea and in plants. It catalyzes the transamination of glutamate-1-semialdehyde to 5-aminolevulinic acid (ALA) in a pyridoxal 5'-phosphate (PLP)-dependent manner. We present the crystal structure of GSAM from the thermophilic cyanobacterium Thermosynechococcus elongatus (GSAM(Tel)) in its PLP-bound form at 2.85A resolution. GSAM(Tel) is a symmetric homodimer, whereas GSAM from Synechococcus (GSAM(Syn)) has been described as asymmetric. The symmetry of GSAM(Tel) thus challenges the previously proposed negative cooperativity between monomers of this enzyme. Furthermore, GSAM(Tel) reveals an extensive flexible region at the interface of the proposed complex of GSAM with glutamyl-tRNA reductase (GluTR), the preceding enzyme in tetrapyrrole biosynthesis. Compared to GSAM(Syn), the monomers of GSAM(Tel) are rotated away from each other along the dimerization interface by 10 degrees . The associated flexibility of GSAM may be essential for complex formation with GluTR to occur. Unexpectedly, we find that GSAM is structurally related to 5-aminolevulinate synthase (ALAS), the ALA-producing enzyme in the Shemin pathway of alpha-proteobacteria and non-plant eukaryotes. This structural relationship applies also to the corresponding subfamilies of PLP-dependent enzymes. We thus propose that the CoA-subfamily (including ALAS) and the aminotransferase subfamily II (including GSAM) are evolutionarily closely related and that ALAS may thus have evolved from GSAM.

5-Aminolevulinate Synthetase↗
PubMed2005

Upregulation of sterol C14-demethylase expression in Trypanosoma cruzi treated with sterol biosynthesis inhibitors.

Infection with the protozoan, Trypanosoma cruzi, is the cause of Chagas disease that occurs widely throughout Latin America. T. cruzi contains sterol biosynthesis enzymes, and produces sterol products similar to those found in fungi. Antifungal drugs that inhibit ergosterol biosynthesis have potent anti-T. cruzi activity in vitro and in animal models. In this report, we describe the effects of sterol biosynthesis inhibitors (simvistatin, zaragosic acid, terbinafine, a lanosterol synthase inhibitor, ketoconazole, and tridemorph) on the regulation of two sterol biosynthesis genes and their protein products. Culturing T. cruzi in the presence of the lanosterol synthase inhibitor, terbinafine, or ketoconazole increased mRNA levels of the sterol C14-demethylase gene approximately 7-12-fold. The sterol C14-demethylase protein levels were also elevated. The effects of the sterol biosynthesis inhibitors on hydroxymethylglutaryl-CoA reductase expression were minimal. Control of the upregulation of sterol C14-demethylase appears to be mediated through the 3'-untranslated region of the gene. The findings demonstrate that T. cruzi can specifically regulate gene expression in response to derangements in its cellular functions.

Animals↗
PubMed2005

An extended signal involved in eukaryotic -1 frameshifting operates through modification of the E site tRNA.

By using a sensitive search program based on hidden Markov models (HMM), we identified 74 viruses carrying frameshift sites among 1500 fully sequenced virus genomes. These viruses are clustered in specific families or genera. Sequence analysis of the frameshift sites identified here, along with previously characterized sites, identified a strong bias toward the two nucleotides 5' of the shifty heptamer signal. Functional analysis in the yeast Saccharomyces cerevisiae demonstrated that high frameshifting efficiency is correlated with the presence of a Psi39 modification in the tRNA present in the E site of the ribosome at the time of frameshifting. These results demonstrate that an extended signal is involved in eukaryotic frameshifting and suggest additional interactions between tRNAs and the ribosome during decoding.

Base Sequence↗