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:3.2.1.31 (beta-glucuronidase)
7,680 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have successfully transferred and expressed a reporter gene driven by an alpha-amylase promoter in a japonica type of rice (Oryza sativa L. cv. Tainung 62) using the Agrobacterium-mediated gene transfer system. Immature rice embryos (10-12 days after anthesis) were infected with an Agrobacterium strain carrying a plasmid containing chimeric genes of beta-glucuronidase (uidA) and neomycin phosphotransferase (nptII). Co-incubation of potato suspension culture (PSC) with the Agrobacterium inoculum significantly improved the transformation efficiency of rice. The uidA and nptII genes, which are under the control of promoters of a rice alpha-amylase gene (alpha Amy8) and Agrobacterium nopaline synthase gene (nos), respectively, were both expressed in G418-resistant calli and transgenic plants. Integration of foreign genes into the genomes of transgenic plants was confirmed by Southern blot analysis. Histochemical localization of GUS activity in one transgenic plant (R0) revealed that the rice alpha-amylase promoter functions in all cell types of the mature leaves, stems, sheaths and roots, but not in the very young leaves. This transgenic plant grew more slowly and produced less seeds than the wild-type plant, but its R1 and R2 progenies grew normally and produced as much seeds as the wild-type plant. Inheritance of foreign genes to the progenies was also confirmed by Southern blot analysis. These data demonstrate successful gene transfer and sexual inheritance of the chimeric genes.
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PMID:Agrobacterium-mediated production of transgenic rice plants expressing a chimeric alpha-amylase promoter/beta-glucuronidase gene. 839 95

A reproducible and efficient transformation system has been developed for maize that is based on direct DNA uptake into embryogenic protoplasts and regeneration of fertile plants from protoplast-derived transgenic callus tissues. Plasmid DNA, containing the beta-glucuronidase (GUS) gene, under the control of the doubled enhancer element (the -208 to -46 bp upstream fragment) from CaMV 35S promoter, linked to the truncated (up to -389 bp from ATG) promoter of wheat, alpha-amylase gene was introduced into protoplasts from suspension culture of HE/89 genotype. The constructed transformation vectors carried either the neomycin phosphotransferase (NPTII) or phosphinothricin acetyltransferase (PAT) gene as selective marker. The applied DNA uptake protocol has resulted at least in 10-20 resistant calli, or GUS-expressing colonies after treatment of 10(6) protoplasts. Vital GUS staining of microcalli has made possible the shoot regeneration from the GUS-stained tissues. 80-90% of kanamycin or PPT resistant calli showed GUS activity, and transgenic plants were regenerated from more than 140 clones. Both Southern hybridization and PCR analysis showed the presence of introduced foreign genes in the genomic DNA of the transformants. The chimeric promoter, composed of a tissue specific monocot promoter, and the viral enhancer element specified similar expression pattern in maize plants, as it was determined by the full CaMV 35S promoter in dicot and other monocot plants. The highest GUS specific activity was found in older leaves with progressively less activity in young leaves, stem and root. Histochemical localization of GUS revealed promoter function in leaf epidermis, mesophyll and vascular bundles, in the cortex and vascular cylinder of the root. In roots, the meristematic tip region and vascular tissues stained intensively. Selected transformants were grown up to maturity, and second-generation seedlings with segregation for GUS activity were obtained after outcrossing. The GUS-expressing segregants carried also the NPTII gene as shown by Southern hybridization.
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PMID:Activity of a chimeric promoter with the doubled CaMV 35S enhancer element in protoplast-derived cells and transgenic plants in maize. 844 39

Functional analysis of a barley high-pI alpha-amylase gene promoter has identified a gibberellin (GA) response complex in the region between -174 and -108. The sequence of the central element, TAACAAA, is very similar to the c-Myb and v-Myb consensus binding site. We investigated the possibility that a GA-regulated Myb transactivates alpha-amylase gene expression in barley aleurone cells. A cDNA clone, GAmyb, which encodes a novel Myb, was isolated from a barley aleurone cDNA library. RNA blot analysis revealed that GAmyb expression in isolated barley aleurone layers is up-regulated by GA. The kinetics of GAmyb expression indicates that it is an early event in GA-regulated gene expression and precedes alpha-amylase gene expression. Cycloheximide blocked alpha-amylase gene expression but failed to block GAmyb gene expression, indicating that protein synthesis is not required for GAmyb gene expression. Gel mobility shift experiments with recombinant GAMyb showed that GAMyb binds specifically to the TAACAAA box in vitro. We demonstrated in transient expression experiments that GAMyb activates transcription of a high-pI alpha-amylase promoter fused to a beta-glucuronidase reporter gene in the absence of GA. Our results indicate that the GAMyb is the sole GA-regulated transcription factor required for transcriptional activation of the high-pI alpha-amylase promoter. We therefore postulate that GAMyb is a part of the GA-response pathway leading to alpha-amylase gene expression in aleurone cells.
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PMID:Gibberellin-regulated expression of a myb gene in barley aleurone cells: evidence for Myb transactivation of a high-pI alpha-amylase gene promoter. 853 41

Knottins are a group of small, disulphide-bonded proteins that bind with high specificity to their target molecules. These proteins appear to use different faces of the protein for their interactions with different targets. Here, we attempted to create knottins with novel binding activities based on the cellulose-binding domain of the fungal enzyme cellobiohydrolase I. Variation was introduced to the face of the protein that binds cellulose. Seven residues, which are located in two regions of the polypeptide chain and form a patch of about 400 A2 on the protein surface, were simultaneously varied by random mutation of the gene. The repertoire was cloned for display on filamentous bacteriophage (5.5 x 10(8) clones), and selected for binding to cellulose or to one of three enzymes (alpha-amylase, alkaline phosphatase and beta-glucuronidase). We thereby isolated variant knottins against cellulose (differing in sequence from the parent knottin) and also against alkaline phosphatase. The binding to (glycosylated) alkaline phosphatase was highly specific with an affinity of about 10 microM, required the presence of disulphide bonds and was mediated through protein (rather than carbohydrate) contacts. Knottin scaffolds therefore appear to be a promising architecture for the creation of small folded proteins with binding activities, with the potential for improvement of binding affinities by mutation, or of using other faces of the protein to provide greater structural diversity in the primary repertoire.
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PMID:Small binding proteins selected from a combinatorial repertoire of knottins displayed on phage. 951 63

The phytohormone abscisic acid (ABA) induces genes-encoding proteins involved in desiccation tolerance and dormancy in seeds, but ABA also suppresses gibberellin (GA)-responsive genes encoding hydrolytic enzymes essential for postgermination growth. A unique serine/threonine protein kinase, PKABA1 mRNA, up-regulated by ABA in seeds, has been identified. In this report, the effect of PKABA1 on the signal transduction pathway mediating ABA induction and suppression of genes has been determined in aleurone layers of barley seeds. Two groups of gene constructs were introduced to barley aleurone layers by using particle bombardment: the reporter constructs containing the coding sequence of beta-glucuronidase gene linked to hormone-responsive promoters and the effector constructs containing the coding region of protein kinases linked to a constitutive promoter. Constitutive expression of PKABA1 drastically suppressed expression of low- and high-pI alpha-amylase and protease genes induced by GA. However, the presence of PKABA1 had only a small effect on the ABA induction of a gene encoding a late embryogenesis abundant protein, HVA1. Our results indicate that PKABA1 acts as a key intermediate in the signal transduction pathway leading to the suppression of GA-inducible gene expression in cereal aleurone layers.
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PMID:An abscisic acid-induced protein kinase, PKABA1, mediates abscisic acid-suppressed gene expression in barley aleurone layers. 999 99

Two Escherichia coli strains in which alpha-amylase production differed were used to study in depth some characteristics related to beta-glucuronidase induction by starch. The beta-glucuronidase background activity in Luria broth medium was comparable for the two isolates, but only amylase positive S1 was able to grow on starch molecules supplied as the sole carbon source. In this case growth resulted at higher beta-glucuronidase levels (p < 0.01) with respect to basal activity and the induced expression was maximal (6.1-fold) when cultures reached the stationary phase. Growth in the presence of a protein synthesis inhibitor (chloramphenicol) was associated with a marked reduction of activity. The beta-glucuronidase activity of amylase negative M94 remained unchanged during starvation on starch medium, but an induced response was observed with methylumbelliferyl-glucuronide. These results further support the hypothesis that starch metabolism is involved in the complex beta-glucuronidase regulation of E. coli strains. This is relevant not only for basic research but also to investigating gut microbial enzymology.
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PMID:Implications of alpha-amylase production and beta-glucuronidase expression in Escherichia coli strains. 1051 Aug 69

The antagonism between gibberellins (GA) and abscisic acid (ABA) is an important factor regulating the developmental transition from embryogenesis to seed germination. In barley aleurone layers, the expression of genes encoding alpha-amylases and proteases is induced by GA but suppressed by ABA. It has been shown that an ABA-induced protein kinase, PKABA1, mediates the ABA suppression of alpha-amylase expression. Using a barley aleurone transient expression system, we have now localized the site of action of PKABA1 relative to other signal transduction components governing the expression of alpha-amylase. The expression of alpha-amylase can be transactivated by the transcription factor GAMyb, which is itself induced by GA. A truncated GAMyb containing the DNA binding domain but lacking the transactivation domain prevents the GA induction of alpha-amylase, further supporting the notion that GAMyb mediates the GA induction of alpha-amylase expression. Although ABA and PKABA1 strongly inhibit the GA induction of alpha-amylase, they have no effect on GAMyb-transactivated alpha-amylase expression. Using a GAMyb promoter--beta-glucuronidase construct, we also show that both ABA and PKABA1 repress the GA induction of GAMyb. In the slender mutant, GAMyb and alpha-amylase are highly expressed, even in the absence of GA. However, this constitutive expression can still be inhibited by ABA, PKABA1, or an inhibitor of cGMP synthesis. On the basis of these observations, we suggest that PKABA1 acts upstream from the formation of functional GAMyb but downstream from the site of action of the Slender gene product. Because PKABA1 inhibits the GA induction of the GAMyb promoter--beta-glucuronidase construct, it appears that at least part of the action of PKABA1 is to downregulate GAMyb at the transcriptional level.
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PMID:Gibberellin/abscisic acid antagonism in barley aleurone cells: site of action of the protein kinase PKABA1 in relation to gibberellin signaling molecules. 1125 Nov 4

Bio-probes that inhibit the action of auxin are useful tools for the study of auxin signaling. To screen for specific inhibitors of auxin signaling, we used an Arabidopsis transgenic line harboring the auxin-inducible promoter derived from PS-IAA4/5 and the reporter gene, GUS (beta-glucuronidase). In this transgenic plant, the exogenous auxin specifically enhanced the expression of the GUS reporter gene. A novel 22-membered spiroketal-macrolide, yokonolide A (1), and related previously known compound, A82548A (2), were isolated from Streptomyces diastatochromogenes B59 as inhibitors of auxin inducible gene expression. The absolute structure of I was determined by detailed spectral analyses and chemical derivatization. 1 and 2 completely inhibited the auxin-induced transcription of the reporter gene at 5 and 1 microm, respectively. In contrast, 1 and 2 did not affect the translation of GUS reporter transcripts. In addition, 1 and 2 did not inhibit the gibberellin-induced alpha-amylase expression at 100 microM in barley aleurone cells. These results suggest that 1 and 2 specifically inhibit auxin signaling leading to auxin-mediated gene expression.
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PMID:Yokonolide A, a new inhibitor of auxin signal transduction, from Streptomyces diastatochromogenes B59. 1156 Mar 76

The in vivo and in vitro effects of 4-amino-3-(D-glucopentitol-1-yl)-5-mercapto-1,2,4-triazole and its 3-methyl analogue on alpha- and beta-glucosidases, beta-glucuronidase as well as alpha-amylase have been investigated. alpha-Glucosidase is the enzyme that is markedly affected in vivo and in vitro in a dose-dependent manner. The compounds showed a reversible inhibition of a competitive type for alpha-glucosidase. Moreover, they exert a relatively potent inhibition on alpha-glucosidase with a Ki magnitude of 3.6 x 10(-4), 9.5 x 10(-5) M.
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PMID:Inhibition of some hepatic glycosidases by the diseco nucleoside, 4-amino-3-(D-glucopentitol-1-yl)-5-mercapto-1,2,4-triazole and its 3-methyl analog. 1250 84

The abscisic acid (ABA) response promoter complexes (ABRCs) of the HVA1 and HVA22 genes have been shown to confer ABA-induced gene expression in cereals. A barley basic domain/Leu zipper (bZIP) transcription factor, HvABI5, is able to recognize ABRCs in vitro in a sequence-specific manner and to transactivate ABRC-beta-glucuronidase reporter genes when introduced to barley aleurone cells via particle bombardment. This transactivation is dependent on the presence of another transcription factor, HvVP1, and cannot be blocked by the negative regulator abi1-1. Using the double-stranded RNA interference technique, we show that HvABI5 and HvVP1 are necessary for the ABA induction of gene expression but have no effect on another hormone-regulated process, the gibberellin-induced and ABA-suppressed expression of alpha-amylase. Our work indicates that although other typical plant bZIP transcription factors may bind ABRCs in vitro, HvABI5 is related to a subfamily of bZIPs responsible for the ABA induction of gene expression. Furthermore, HvABI5 and HvVP1 are not involved in the ABA suppression of gene expression.
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PMID:The transcription factors HvABI5 and HvVP1 are required for the abscisic acid induction of gene expression in barley aleurone cells. 1250 36


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