Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Characterization of CalE10, the N-oxidase involved in calicheamicin hydroxyaminosugar formation.

ABSTRACT: As the first in vitro characterization of a sugar N-oxidase, this study establishes CalE10 as the key oxidase involved in calicheamicin hydroxylamino glycoside formation. This study confirms that oxidation occurs at the sugar nucleotide stage prior to glycosyltransfer, and substrate specificity studies reveal CalE10-catalyzed oxidation to be regiospecific and to present trace amounts of the corresponding nitrosugar in vitro. This work also sets a precedent for the future study of other N-oxidases involved in hydroxylamino-, nitroso-, and/or nitrosugar formation.

SUBMITTER: Johnson HD

PROVIDER: S-EPMC2704561 | biostudies-literature | 2008 Dec

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

Characterization of CalE10, the N-oxidase involved in calicheamicin hydroxyaminosugar formation.

Johnson Heather D HD Thorson Jon S JS

Journal of the American Chemical Society 20081201 52

As the first in vitro characterization of a sugar N-oxidase, this study establishes CalE10 as the key oxidase involved in calicheamicin hydroxylamino glycoside formation. This study confirms that oxidation occurs at the sugar nucleotide stage prior to glycosyltransfer, and substrate specificity studies reveal CalE10-catalyzed oxidation to be regiospecific and to present trace amounts of the corresponding nitrosugar in vitro. This work also sets a precedent for the future study of other N-oxidase ...[more]

PMID: 19055330

Similar Datasets

Structural characterization of CalO2: a putative orsellinic acid P450 oxidase in the calicheamicin biosynthetic pathway.

Project description:Although bacterial iterative Type I polyketide synthases are now known to participate in the biosynthesis of a small set of diverse natural products, the subsequent downstream modification of the resulting polyketide products remains poorly understood. Toward this goal, we report the X-ray structure determination at 2.5 A resolution and preliminary characterization of the putative orsellenic acid P450 oxidase (CalO2) involved in calicheamicin biosynthesis. These studies represent the first crystal structure for a P450 involved in modifying a bacterial iterative Type I polyketide product and suggest the CalO2-catalyzed step may occur after CalO3-catalyzed iodination and may also require a coenzyme A- (CoA) or acyl carrier protein- (ACP) bound substrate. Docking studies also reveal a putative docking site within CalO2 for the CLM orsellinic acid synthase (CalO5) ACP domain which involves a well-ordered helix along the CalO2 active site cavity that is unique compared with other P450 structures.

| S-EPMC2742692 | biostudies-literature

Structural Characterization of CalS8, a TDP-α-D-Glucose Dehydrogenase Involved in Calicheamicin Aminodideoxypentose Biosynthesis.

Project description:Classical UDP-glucose 6-dehydrogenases (UGDHs; EC 1.1.1.22) catalyze the conversion of UDP-α-d-glucose (UDP-Glc) to the key metabolic precursor UDP-α-d-glucuronic acid (UDP-GlcA) and display specificity for UDP-Glc. The fundamental biochemical and structural study of the UGDH homolog CalS8 encoded by the calicheamicin biosynthetic gene is reported and represents one of the first studies of a UGDH homolog involved in secondary metabolism. The corresponding biochemical characterization of CalS8 reveals CalS8 as one of the first characterized base-permissive UGDH homologs with a >15-fold preference for TDP-Glc over UDP-Glc. The corresponding structure elucidations of apo-CalS8 and the CalS8·substrate·cofactor ternary complex (at 2.47 and 1.95 Å resolution, respectively) highlight a notably high degree of conservation between CalS8 and classical UGDHs where structural divergence within the intersubunit loop structure likely contributes to the CalS8 base permissivity. As such, this study begins to provide a putative blueprint for base specificity among sugar nucleotide-dependent dehydrogenases and, in conjunction with prior studies on the base specificity of the calicheamicin aminopentosyltransferase CalG4, provides growing support for the calicheamicin aminopentose pathway as a TDP-sugar-dependent process.

| S-EPMC4646273 | biostudies-literature

Structural and functional characterization of CalS11, a TDP-rhamnose 3'-O-methyltransferase involved in calicheamicin biosynthesis.

Project description:Sugar methyltransferases (MTs) are an important class of tailoring enzymes that catalyze the transfer of a methyl group from S-adenosyl-l-methionine to sugar-based N-, C- and O-nucleophiles. While sugar N- and C-MTs involved in natural product biosynthesis have been found to act on sugar nucleotide substrates prior to a subsequent glycosyltransferase reaction, corresponding sugar O-methylation reactions studied thus far occur after the glycosyltransfer reaction. Herein we report the first in vitro characterization using (1)H-(13)C-gHSQC with isotopically labeled substrates and the X-ray structure determination at 1.55 Å resolution of the TDP-3'-O-rhamnose-methyltransferase CalS11 from Micromonospora echinospora. This study highlights a unique NMR-based methyltransferase assay, implicates CalS11 to be a metal- and general acid/base-dependent O-methyltransferase, and as a first crystal structure for a TDP-hexose-O-methyltransferase, presents a new template for mechanistic studies and/or engineering.

| S-EPMC3875630 | biostudies-literature

Characterization of the calicheamicin orsellinate C2-O-methyltransferase CalO6.

Project description:Although bacterial iterative type I polyketide synthases are now known to participate in the biosynthesis of a small set of diverse natural products, the subsequent downstream modification of the resulting polyketide products is poorly understood. We report the functional characterization of the putative orsellinic acid C2-O-methyltransferase, which is involved in calicheamicin biosynthesis. This study suggests that C2-O-methylation precedes C3-hydroxylation/methylation and C5-iodination and requires a coenzyme A- or acyl carrier protein-bound substrate.

| S-EPMC4111469 | biostudies-literature

Structural characterization of EasH (Aspergillus japonicus) - an oxidase involved in cycloclavine biosynthesis.

Project description:Aj_EasH is a non-heme iron- and α-keto-glutarate-dependent oxidase that is responsible for an unusual cyclopropyl ring formation in the biosynthesis of the fungal ergot alkaloid cycloclavine. The three dimensional structure of Aj_EasH (2.2 Å resolution) reported here provides insight into the mechanism of this unusual and complex reaction.

| S-EPMC5317212 | biostudies-literature

Hypoxia-inducible factor 1-regulated lysyl oxidase is involved in Staphylococcus aureus abscess formation.

Project description:Hypoxia-inducible factor 1 (HIF-1) is the key transcription factor involved in the adaptation of mammals to hypoxia and plays a crucial role in cancer angiogenesis. Recent evidence suggests a leading role for HIF-1 in various inflammatory and infectious diseases. Here we describe the role of HIF-1 in Staphylococcus aureus infections by investigating the HIF-1-dependent host cell response. For this purpose, transcriptional profiling of HIF-1α-deficient HepG2 and control cells, both infected with Staphylococcus aureus, was performed. Four hours after infection, the expression of 190 genes, 24 of which were regulated via HIF-1, was influenced. LOX (encoding lysyl oxidase) was one of the upregulated genes with a potential impact on the course of S. aureus infection. LOX is an amine oxidase required for biosynthetic cross-linking of extracellular matrix components. LOX was upregulated in vitro in different cell cultures infected with S. aureus and also in vivo, in kidney abscesses of mice intravenously infected with S. aureus and in clinical skin samples from patients with S. aureus infections. Inhibition of LOX by β-aminopropionitrile (BAPN) did not affect the bacterial load in kidneys or blood but significantly influenced abscess morphology and collagenization. Our data provide evidence for a crucial role of HIF-1-regulated LOX in abscess formation.

| S-EPMC3697610 | biostudies-literature

Structural characterization of CalO1: a putative orsellinic acid methyltransferase in the calicheamicin-biosynthetic pathway.

Project description:The X-ray structure determination at 2.4?Å resolution of the putative orsellinic acid C3 O-methyltransferase (CalO1) involved in calicheamicin biosynthesis is reported. Comparison of CalO1 with a homology model of the functionally related calicheamicin orsellinic acid C2 O-methyltransferase (CalO6) implicates several residues that are likely to contribute to the regiospecificity of alkylation. Consistent with the proposed requirement of an acyl-carrier-protein-bound substrate, this structural study also reveals structural determinants within CalO1 that are anticipated to accommodate an association with an acyl carrier protein.

| S-EPMC3046457 | biostudies-literature

Characterization of a novel oxidase from Thelonectria discophora SANK 18292 involved in nectrisine biosynthesis.

Project description:A fungus, Thelonectria discophora SANK 18292 (JCM 30947), produces nectrisine that has a nitrogen-containing heterocyclic 5-membered ring acting as a glycosidase inhibitor. Our previous study showed the possibility that 4-amino-4-deoxyarabinitol was enzymatically converted to nectrisine but the enzyme was not known. In order to characterize the enzyme, which is designated as NecC, it was purified from the fungus using ammonium sulfate precipitation and anion exchange chromatography. Liquid chromatography-tandem mass spectrometry analysis of NecC tryptic digests revealed partial NecC protein sequences. Subsequently, the partial DNA fragments were amplified by polymerase chain reaction with degenerate oligonucleotide primers and cloned. Then, necC complete genomic DNA was cloned by screening a genomic library of the fungus. Recombinant NecC also had NecC enzymatic activity, thus providing verification for the necC gene. NecC presumably belonged to the family of glucose methanol choline oxidoreductases, forming oligomers ranging approximately from 8 mer to 16 mer based on the results of native PAGE, and was also found to have a melting temperature of 57 °C, an optimal reaction condition of pH 7 at 30 °C, an activity inhibited by Cu(2+) or ethylenediaminetetraacetic acid, and 4-amino-4-deoxyarabinitol as its preferred substrate. It was also indicated that not nectrisine but 4-amino-4-deoxyarabinitol was mainly extracted from the mycelium, and then was converted to nectrisine by the enzyme NecC in vitro. We believe that these findings are helpful to establish a nectrisine manufacturing process at large scale with the fungus.

| S-EPMC4718913 | biostudies-literature

fbfB, a gene encoding a putative galactose oxidase, is involved in Stigmatella aurantiaca fruiting body formation.

Project description:Stigmatella aurantiaca is a gram-negative bacterium which forms, under conditions of starvation in a multicellular process, characteristic three-dimensional structures: the fruiting bodies. For studying this complex process, mutants impaired in fruiting body formation have been induced by transposon insertion with a Tn5-derived transposon. The gene affected (fbfB) in one of the mutants (AP182) was studied further. Inactivation of fbfB results in mutants which form only clumps during starvation instead of wild-type fruiting bodies. This mutant phenotype can be partially rescued, if cells of mutants impaired in fbfB function are mixed with those of some independent mutants defective in fruiting before starvation. The fbfB gene is expressed about 14 h after induction of fruiting body formation as determined by measuring beta-galactosidase activity in a merodiploid strain harboring the wild-type gene and an fbfB-delta trp-lacZ fusion gene or by Northern (RNA) analysis with the Rhodobacter capsulatus pufBA fragment fused to fbfB as an indicator. The predicted polypeptide FbfB has a molecular mass of 57.8 kDa and shows a significant homology to the galactose oxidase (GaoA) of the fungus Dactylium dendroides. Galactose oxidase catalyzes the oxidation of galactose and primary alcohols to the corresponding aldehydes.

| S-EPMC107013 | biostudies-literature

Characterization of Aldehyde Oxidase (AO) Genes Involved in the Accumulation of Carotenoid Pigments in Wheat Grain.

Project description:Aldehyde Oxidase (AO) enzyme (EC 1.2.3.1) catalyzes the final steps of carotenoid catabolism and it is a key enzyme in the abscisic acid (ABA) biosynthesis. AO isoforms are located in the cytosolic compartment of tissues in many plants, where induce the oxidation of aldehydes into carboxylic acid, and in addition, catalyze the hydroxylation of some heterocycles. The goal of the present study was to characterize the AO genes involved in the accumulation of carotenoid pigments in wheat grain, an important quantitative trait controlled by multiple genes. The cDNAs corresponding to the four AO isoforms from Arabidopsis thaliana and five AO isoforms from Brachypodium distachyon were used as query in 454 sequence assemblies data for Triticum aestivum cv. Chinese Spring (https://urgi.versailles.inra.fr/blast/blast.php) to obtain the partial or whole orthologous wheat AO sequences. Three wheat isoforms, designated AO1, AO2, and AO3 were located on the chromosome groups 2, 5, and 7, respectively, and mapped on two consensus wheat maps by SNP markers located within the AO gene sequences. To validate the possible relationships between AO3 genes and carotenoid accumulation in wheat, the expression levels of AO-A3 and AO-B3 gene were determined during the kernel maturation stage of two durum wheat cultivars, Ciccio and Svevo, characterized by a low and high carotenoid content, respectively. Different AO-A3 gene expression values were observed between the two cultivars indicating that the AO-A3 allele present in Ciccio was more active in carotenoid degradation. A gene marker was developed and can be used for marker-assisted selection in wheat breeding programs.

| S-EPMC5443152 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data