Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.2.1.31 (beta-glucuronidase)
 7,680 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The RNA genome of tobacco etch potyvirus (TEV) was engineered to express bacterial beta-glucuronidase (GUS) fused to the virus helper component proteinase (HC-Pro). It was shown previously that prolonged periods (approximately 1 month) of TEV-GUS propagation in plants resulted in the appearance of spontaneous deletion variants. Nine deletion mutants were identified by nucleotide sequence analysis of 40 cDNA clones obtained after polymerase chain reaction amplification. The mutants were missing between 1,741 and 2,074 nucleotides from TEV-GUS, including the sequences coding for most of GUS and the N-terminal region of HC-Pro. This region of HC-Pro contains determinants involved in helper component activity during aphid transmission, as well as a highly conserved series of cysteine residues. The deletion variants were shown to replicate and move systemically without the aid of a helper virus. Infectious viruses harboring the two largest HC-Pro deletions (termed TEV-2del and TEV-7del) were reconstructed by subcloning the corresponding mutated regions into full-length DNA copies of the TEV genome. Characterization of these and additional variants derived by site-directed mutagenesis demonstrated that deletion of sequences coding for the HC-Pro N-terminal domain had a negative effect on accumulation of viral RNA and coat protein. The TEV-2del variant possessed an aphid-nontransmissible phenotype that could be rescued partially by prefeeding of aphids on active HC-Pro from another potyvirus. These data suggest that the N-terminal domain of HC-Pro or its coding sequence enhances virus replication or genome expression but does not provide an activity essential for these processes. The function of this domain, as well as a proposed deletion mechanism involving nonhomologous recombination, is discussed.
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PMID:Spontaneous mutagenesis of a plant potyvirus genome after insertion of a foreign gene. 837 51

Superoxide dismutases (SODs; superoxide: superoxide oxidoreductase, EC 1.15.1.1) play a key role in protection against oxygen radicals, and SOD gene expression is highly induced during environmental stress. To determine the conditions of SOD induction, the promoter of the cytosolic copper/zinc SOD (Cu/ZnSODcyt) gene was isolated in Nicotiana plumbaginifolia and fused to the beta-glucuronidase reporter gene. Oxidative stress is likely to alter the cellular redox in favor of the oxidized status. Surprisingly, the expression of the Cu/ZnSODcyt gene is induced by sulfhydryl antioxidants such as reduced glutathione, cysteine, and dithiothreitol, whereas the oxidized forms of glutathione and cysteine have no effect. It is therefore possible that reduced glutathione directly acts as an antioxidant and simultaneously activates the Cu/ZnSODcyt gene during oxidative stress.
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PMID:Redox-activated expression of the cytosolic copper/zinc superoxide dismutase gene in Nicotiana. 846 30

The pharmacodynamics of the non-steroidal anti-inflammatory drugs flunixin, tolfenamic acid and ketoprofen were studied in calves after intravenous administration. An acute inflammatory reaction was induced in tissue cages by the intracaveal injection of the mild irritant carrageenan, and the inhibition of inflammatory mediators and enzymes was investigated. The substances measured in the exudate included the enzymes (active and total metalloproteases, serine and cysteine proteases, acid phosphatase [AP], lactate dehydrogenase [LDH] and beta-glucuronidase) and the eicosanoids (prostaglandin [PG]E2 and leukotriene [LT]B4). Studies were also made of inhibition of the synthesis of serum thromboxane (Tx)B2 ex vivo, of bradykinin-induced oedema in vivo and of the generation of superoxide anions (O2-) in vitro. None of the drugs affected the concentration of LTB4, or the activities of metalloproteases, cysteine and serine proteases, AP or LDH in the exudate. All the drugs inhibited the synthesis of serum TxB2 and exudate PGE2 and inhibited the release of beta-glucuronidase. They also decreased the oedematous response to intradermally injected bradykinin and inhibited the generation of O2- ions by neutrophils in vitro. These actions may contribute to the anti-inflammatory effects of the drugs and hence to their clinical efficacy.
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PMID:Comparative pharmacodynamics of flunixin, ketoprofen and tolfenamic acid in calves. 856 Jul 1

The barley cysteine proteinase B (EPB) is the main protease responsible for the degradation of endosperm storage proteins providing nitrogenous nutrients to support the growth of young seedlings. The expression of this enzyme is induced in the germinating seeds by the phytohormone, gibberellin, and suppressed by another phytohormone, abscisic acid. In situ hybridization experiments indicate that EPB is expressed in the scutellar epithelium within 24 h of seed germination, but the aleurone tissue surrounding the starchy endosperm eventually becomes the main tissue expressing this enzyme. The EPB gene family of barley consists of two very similar genes, EPB1 and EPB2, both of which have been mapped to chromosome 3. The sequences of EPB1 and EPB2 match with the two previously published cDNA clones indicating that both genes are expressed. Interestingly, neither of these genes contain any introns, a rare phenomenon in which all members of a small gene family are active intronless genes. Sequence comparison indicates that the barley EPB family can be classified as cathepsin L-like endopeptidases and is most closely related to two legume cysteine proteinases (Phaseolus vulgaris EP-C1 and Vigna mungo SHEP) which are also involved in seed storage protein degradation. The promoters of EPB1 and EPB2 have been linked to the coding sequence of a reporter gene, GUS, encoding beta-glucuronidase, and introduced into barley aleurone cells using the particle bombardment method. Transient expression studies indicate that EPB promoters are sufficient to confer the hormonal regulation of these genes.
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PMID:A major cysteine proteinase, EPB, in germinating barley seeds: structure of two intronless genes and regulation of expression. 875 90

We have characterized cotton (Gossypium hirsutum L.) genes encoding type 1 metallothionein-like proteins that are highly expressed in roots. Little or no expression of these genes was detected in other organs and tissues. The deduced amino acid sequences have a high degree of similarity with type 1 metallothionein-like proteins from other plants, including a central hydrophobic domain flanked by conserved cysteine-rich motifs. The type 1 metallothionein-like genes of cotton are encoded by a small gene family. One gene (MT1-A) was analyzed in detail and found to have three exons which are 52, 83 and 397 bp long, and two introns 130 and 1042 bp in length. Three of the type 1 metallothionein-like genes are organized in a tandom array, and the 5'-flanking regions of these genes share a high degree of sequence similarity. Two of the clustered genes (MT1-A and MT1-B) are expressed at about equal levels in roots and use the same transcription start site. A 640 bp promoter fragment from the MT1-A gene was sufficient to direct expression of beta-glucuronidase (GUS) in transformed cotton roots. The expression was highest near the root tip.
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PMID:Characterization and expression of metallothionein-like genes in cotton. 879 Mar 3

Conjugation of multiple ubiquitins serves as a committed step in the degradation of a variety of intracellular eukaryotic proteins by the 26S proteasome. Conjugates are formed via a three-enzyme cascade; the initial step requires ubiquitin-activating enzyme (E1), which couples ubiquitin activation to ATP hydrolysis. Previously, we showed that many higher plants contain multiple E1 proteins and described several E1 genes from wheat. To facilitate understanding of the roles of the different plant E1s, we characterized the E1 gene and protein family from Arabidopsis thaliana. Arabidopsis E1s are encoded by two genes (AtUBA1 and AtUBA2) that synthesize approximately 123-kDa proteins with 81% amino acid sequence identity to each other and 44-75% sequence identity with confirmed E1s from other organisms. Like other E1 proteins, AtUBA1 and 2 contain a cysteine residue in the putative active site for forming the ubiquitin thiol-ester intermediate. Enzymatic analysis of the corresponding proteins expressed in Escherichia coli demonstrated that both proteins activate ubiquitin in an ATP-dependent reaction and transfer the activated ubiquitin to a variety of Arabidopsis E2s with near equal specificity. Expression studies by quantitative RT-PCR and histochemistry with transgenic plants containing AtUBA promoter-beta-glucuronidase-coding region fusions showed that the AtUBA1 and 2 genes are co-expressed in most, if not all, Arabidopsis tissues and cells. Collectively, the data indicate that E1 proteins, and presumably the rest of the ubiquitin pathway, are present throughout Arabidopsis. They also show that the AtUBA1 and 2 genes are not differentially expressed nor do they encode E1s with dramatically distinct enzymatic properties.
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PMID:The ubiquitin-activating enzyme (E1) gene family in Arabidopsis thaliana. 907 89

Synchronously dividing cell cultures of Catharanthus roseus were used to isolate cDNAs for two mitotic cyclins, named CYS and CYM. The deduced protein sequence of CYS is similar to that of A-type cyclins, and CYM belongs to the group of B-type cyclins. In a fashion similar to the pattern of expression seen for A-type and B-type cyclins in mammalian cells, CYS is expressed before CYM in C. roseus cells during the cell cycle. CYS mRNA accumulated at the onset of S phase and disappeared early in the G2 phase, whereas CYM mRNA was detected in the G2 and M phases of the cell cycle. Tobacco homologs of the two genes showed similar cell-cycle dependent expression patterns in synchronous cultures of tobacco BY2 cells. In both systems, CYS was expressed much earlier in the cell cycle than most other plant A-type cyclins, and hence CYS along with the soybean cyc1GM can be classified into a distinct subclass. The activities of CYM and CYS promoters during the cell cycle were analyzed in stably transformed tobacco BY2 cells. Cyclin promoter sequences of 0.5 kb could confer the typical cell-cycle-dependent expression to the beta-glucuronidase (GUS) reporter gene: the CYS promoter directed S-phase-specific expression, whereas the CYM promoter drove M-phase-specific expression. These results indicate the important role of transcriptional regulation in the oscillations of cyclin mRNA levels during the cell cycle.
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PMID:Cell-cycle-regulated transcription of A- and B-type plant cyclin genes in synchronous cultures. 919 70

An assay was developed for the simultaneous measurement of cyclohexene oxide and its metabolites (cyclohexanol, trans-cyclohexane-1,2-diol, cyclohexane-1,2-diol-O-glucuronide, and N-acetyl-S-(2-hydroxycyclohexyl)-L-cysteine) in rat urine and plasma using gas chromatography. A mixture of ethyl acetate-acetonitrile (70:30) was used as the extracting solvent for both matrices. This liquid-liquid extraction procedure was followed by the separation of cyclohexene oxide and its metabolites on an HP-FFAP fused-silica capillary column. In order to determine the amount of cyclohexane-1,2-diol-O-glucuronide, samples were incubated at 37 degrees C with beta-glucuronidase and the amount of cyclohexane-1,2-diol formed from the reaction determined. The extraction efficiencies of cyclohexene oxide and cyclohexanol were greater than 90% both in urine and plasma. However, recovery from the plasma and urine for trans-cyclohexane-1,2-diol (60-68%) and N-acetyl-S-(2-hydroxycyclohexyl)-L-cysteine (approximately 76%) were considerably less. Long term stability studies showed that urine samples spiked with cyclohexene oxide and trans-cyclohexane-1,2-diol are stable at -20 degrees C for up to 9 weeks. However, plasma samples are only stable for up to 2 weeks under the same conditions. The calibration curves for all analytes were linear over the range of 12.5 to 400 micrograms/ml and correlation coefficients (r2) were greater than 0.990. The limit of detection for cyclohexene oxide, cyclohexanol, and N-acetyl-S-(2-hydroxycyclohexyl)-L-cysteine is 1.56 micrograms/ml, while the limit of detection for trans-cyclohexane-1,2-diol is 3.12 micrograms/ml. This method has been used for the determination of the disposition and metabolism of cyclohexene oxide, and may be applied in environmental monitoring, as well as in microbiological studies for other epoxide materials.
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PMID:Simultaneous determination of cyclohexene oxide and its metabolites in rat plasma and urine by gas chromatography. 930 Sep 9

Biomarkers of periodontal disease activity may be obtained from potential proteolytic and hydrolytic enzymes of inflammatory cell origin. Studies that have sought to correlate these enzymes with periodontal disease activity are reviewed with special consideration given to collagenases, cysteine, aspartate and serine proteinases, beta-glucuronidase, arylsulphate, alkaline and acid phosphatases, myeloperoxidase, lysozyme and lactoferrin.
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PMID:Advances in periodontal diagnosis. 7. Proteolytic and hydrolytic enzymes link with periodontitis. 959 84

The phenomenon of enantioselectivity in the metabolism of mexiletine (MEX) conjugation was investigated in eight female patients with the arrhythmic form of chronic Chagas' heart disease treated with racemic mexiletine hydrochloride (two 100 mg capsules every 8 hr). Blood samples were collected up to 24 hr after the administration of the morning dose, with discontinuation of the subsequent doses during the study period. Plasma concentrations of N-hydroxymexiletine glucuronide were calculated as the difference between the concentrations of unchanged and total (unchanged + conjugated) MEX enantiomers. Total plasma MEX concentrations were analyzed by HPLC after enzymatic hydrolysis with beta-glucuronidase, the formation of diastereomeric derivatives with the chiral reagent N-acetyl-L-cysteine/o-phthalaldehyde, and fluorescence detection. The differences in the pharmacokinetic parameters of the enantiomers were evaluated by the paired t-test. The plasma concentrations of the (+)-(S)-MEX did not differ before and after enzymatic hydrolysis. The pharmacokinetic parameters calculated for (-)-(R)-N-hydroxymexiletine glucuronide are presented as means (95% confidence interval): maximum plasma concentration Cmax = 194.0 ng.ml-1 (154.3-233.7), time to maximum plasma concentration tmax = 1.4 hr (0.3-2.5), area under the plasma concentration versus time curve AUC0-24 = 2099.2 ng.h.ml-1 (1585.6-2612.6), elimination half-life t1/2 beta = 12.8 hr (9.9-15.6) and extent of conjugation of 31.6% (24.3-38.9%). The present data indicate stereospecific conjugation of (-)-(R)-N-hydroxymexiletine in the female patients with the arrhythmic form of Chagas' heart disease.
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PMID:Enantioselectivity in the metabolism of mexiletine by conjugation in female patients with the arrhythmic form of chronic Chagas' heart disease. 991 50


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