Search results for "Glucosyltransferase"
showing 10 items of 32 documents
Separation by FPLC chromatofocusing of UDP-glucosyltransferases from three developmental stages of Drosophila melanogaster.
1997
Variation of UDP-glucosyltransferase activity, during Drosophila melanogaster development, was analyzed. The endogenous metabolite xanthurenic acid and the xenobiotic compounds 1-naphthol and 2-naphthol were used as substrates. Developmentally regulated differences were observed for the three substrates, suggesting the presence of UDP-glucosyltransferase isoenzymes. This was further confirmed by FPLC chromatofocusing on a Mono P column: seven peaks of UDP-glucosyltransferase activity (pHs: ≥6.3, 5.8, 5.5, 4.9, 4.5, 4.2, ≤4.0) with either single or overlapping substrate specificity were detected. A single xanthurenic acid:UDP-glucosyltransferase activity (pl 5.8) was found throughout develop…
Hydroquinone: O-glucosyltransferase from cultivated Rauvolfia cells: enrichment and partial amino acid sequences.
2000
Plant cell suspension cultures of Rauvolfia are able to produce a high amount of arbutin by glucosylation of exogenously added hydroquinone. A four step purification procedure using anion exchange, hydrophobic interaction, hydroxyapatite-chromatography and chromatofocusing delivered in a yield of 0.5%, an approximately 390 fold enrichment of the involved glucosyltransferase. SDS-PAGE showed a M(r) for the enzyme of 52 kDa. Proteolysis of the pure enzyme with endoproteinase LysC revealed six peptide fragments with 9-23 amino acids which were sequenced. Sequence alignment of the six peptides showed high homologies to glycosyltransferases from other higher plants.
Enhanced Accumulation of Betulinic Acid in Transgenic Hairy Roots of Senna obtusifolia Growing in the Sprinkle Bioreactor and Evaluation of Their Bio…
2021
Betulinic acid, which is found in transgenic roots of Senna obtusifolia (L.) H.S.Irwin & Barneby, is a pentacyclic triterpene with distinctive pharmacological activities. In this study, we report the differences in the content of betulinic acid and selected anthraquinones in transgenic S. obtusifolia hairy roots with overexpression of the PgSS1 gene (SOPSS2 line) and in transformed hairy roots without this genetic construct (SOA41 line). Both hairy root lines grew in 10 L sprinkle bioreactor. Additionally, the extracts obtained from this plant material were used for biological tests. Our results demonstrated that the SOPSS2 hairy root cultures from the bioreactor showed an increase in the c…
Inhibition of FcεRI-mediated Activation of Rat Basophilic Leukemia Cells by Clostridium difficile Toxin B (Monoglucosyltransferase)
1996
Abstract Treatment of rat basophilic leukemia (RBL) 2H3-hm1 cells with Clostridium difficile toxin B (2 ng/ml), which reportedly depolymerizes the actin cytoskeleton, blocked [3H]serotonin release induced by 2,4-dinitrophenyl-bovine serum albumin, carbachol, mastoparan, and reduced ionophore A23187-stimulated degranulation by about 55-60%. In lysates of RBL cells, toxin B 14C-glucosylated two major and one minor protein. By using two-dimensional gel electrophoresis and immunoblotting, RhoA and Cdc42 were identified as protein substrates of toxin B. In contrast to toxin B, Clostridium botulinum transferase C3 that selectively inactivates RhoA by ADP-ribosylation did not inhibit degranulation…
Spatial analysis of plant metabolism: Sucrose imaging within Vicia faba cotyledons reveals specific developmental patterns
2002
During legume embryogenesis the differentiation of the cotyledons proceeds gradually in a wave-like manner. The process is metabolically and genetically controlled and regulated by sugars. In order to perform a spatial and temporal analysis of the sugar distribution pattern a new method was developed to specifically measure sucrose directly in tissues via bioluminescence and single photon counting. This enabled a quantitative sucrose imaging with a resolution close to the single cell level. The procedure was applied on sections of Vicia faba cotyledons covering the main stages of histodifferentiation. Young embryos before the storage phase contained moderate levels of sucrose, which were ev…
Glucosylation of Rho proteins by Clostridium difficile toxin B.
1995
TOXIN A and B, the major virulence factors of Clostridium difficile, are the causative agents of antibiotic-associated pseudomembran-ous colitis. In cultured cell lines their potent cytotoxicity results from their ability to induce disaggregation of the microfilament cytoskeleton1,2. Toxin B acts on the low-molecular-mass GTPase Rho A3,4, which is involved in the regulation of the actin cytoskeleton. We report here that toxin B catalyses the incorporation of up to one mole of glucose per mole of RhoA at the amino acid thre-onine at position 37. The modification was identified and localized by tandem electrospray mass spectrometry. UDP-glucose selectively serves as cosubstrate for the monogl…
Ras, Rap, and Rac Small GTP-binding Proteins Are Targets for Clostridium sordellii Lethal Toxin Glucosylation
1996
Lethal toxin (LT) from Clostridium sordellii is one of the high molecular mass clostridial cytotoxins. On cultured cells, it causes a rounding of cell bodies and a disruption of actin stress fibers. We demonstrate that LT is a glucosyltransferase that uses UDP-Glc as a cofactor to covalently modify 21-kDa proteins both in vitro and in vivo. LT glucosylates Ras, Rap, and Rac. In Ras, threonine at position 35 was identified as the target amino acid glucosylated by LT. Other related members of the Ras GTPase superfamily, including RhoA, Cdc42, and Rab6, were not modified by LT. Incubation of serum-starved Swiss 3T3 cells with LT prevents the epidermal growth factor-induced phosphorylation of m…
Metazoan Circadian Rhythm: Toward an Understanding of a Light-Based Zeitgeber in Sponges
2013
In all eukaryotes, the 24-h periodicity in the environment contributed to the evolution of the molecular circadian clock. We studied some elements of a postulated circadian clock circuit in the lowest metazoans, the siliceous sponges. First, we identified in the demosponge Suberites domuncula the enzyme luciferase that generates photons. Then (most likely), the photons generated by luciferase are transmitted via the biosilica glass skeleton of the sponges and are finally harvested by cryptochrome in the same individual; hence, cryptochrome is acting as a photosensor. This information-transduction system, generation of light (luciferase), photon transmission (through the siliceous spicules),…
UDP-glucose deficiency in a mutant cell line protects against glucosyltransferase toxins from Clostridium difficile and Clostridium sordellii.
1996
Abstract We have previously isolated a fibroblast mutant cell with high resistance to the two Rho-modifying glucosyltransferase toxins A and B of Clostridium difficile. We demonstrate here a low level of UDP-glucose in the mutant, which explains its toxin resistance since: (i) to obtain a detectable toxin B-mediated Rho modification in lysates of mutant cells, addition of UDP-glucose was required, and it promoted the Rho modification dose-dependently; (ii) high pressure liquid chromatography analysis of nucleotide extracts of cells indicated that the level of UDP-glucose in the mutant (0.8 nmol/106 cells) was lower than in the wild type (3.7 nmol/106 cells); and (iii) sensitivity to toxin B…
Utilizing genetically engineered bacteria to produce plant-specific glucosides
2001
Plant-derived glucosides have attracted much attention due to their widespread applications. This class of products is difficult to isolate or to synthesize in pure form because of the resulting low yields. Thus, simple approaches for the generation of such glucosides would be highly beneficial. We purified and characterized a novel glucosyltransferase from plant cell suspension cultures of Rauvolfia serpentina, which showed rather low substrate specificity. We obtained its cDNA and expressed the active recombinant protein in bacteria (Escherichia coli) with excellent plant-specific glucosylation efficiencies. Compared with the plant system, the bacteria delivered the new enzyme, which was …