Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.30.1 (S1 nuclease)
 3,660 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The DNA sequence of the pea cytosolic glutamine synthetase GS3A gene promoter has been determined and the start of transcription mapped using S1 nuclease. The full-length promoter and a series of 5' deletions were fused to beta-glucuronidase (GUS) and introduced into transgenic tobacco and alfalfa. In transgenic tobacco the GS3A promoter directed GUS expression in the phloem cells of the vasculature in leaves, stems and roots. GUS expression was also detected in the vasculature of cotyledons and the root tips of germinating T1 seedlings. The promoter conferred a similar expression pattern in transgenic alfalfa, and expression was also observed in root nodules. Nodule expression was located in nodule primordia, as well as the meristem, symbiotic zone, and vasculature of mature nodules. The promoter was found to be active even when deleted to -132 relative to the start of transcription. DNA mobility-shift analysis identified a protein present in nuclear and whole-cell plant extracts which bound to a 17 bp DNA element contained within the minimal -132 promoter required for expression.
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PMID:A promoter sequence involved in cell-specific expression of the pea glutamine synthetase GS3A gene in organs of transgenic tobacco and alfalfa. 168 48

We have isolated the Bradyrhizobium japonicum gene encoding glutamine synthetase I (glnA) from a phage lambda library by using a fragment of the Escherichia coli glnA gene as a hybridization probe. The rhizobial glnA gene has homology to the E. coli glnA gene throughout the entire length of the gene and can complement an E. coli glnA mutant when borne on an expression plasmid in the proper orientation to be transcribed from the E. coli lac promoter. High levels of glutamine synthetase activity can be detected in cell-free extracts of the complemented E. coli. The enzyme encoded by the rhizobial gene was identified as glutamine synthetase I on the basis of its sedimentation properties and resistance to heat inactivation. DNA sequence analysis predicts a high level of amino acid sequence homology among the amino termini of B. japonicum, E. coli, and Anabaena sp. strain 7120 glutamine synthetases. S1 nuclease protection mapping indicates that the rhizobial gene is transcribed from a single promoter 131 +/- 2 base pairs upstream from the initiation codon. This glnA promoter is active when B. japonicum is grown both symbiotically and in culture with a variety of nitrogen and carbon sources. There is no detectable sequence homology between the constitutively expressed glnA promoter and the differentially regulated nif promoters of the same B. japonicum strain.
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PMID:Characterization of the gene encoding glutamine synthetase I (glnA) from Bradyrhizobium japonicum. 285 70

A human genomic clone, psi GS, containing an intron-less glutamine synthetase (GS)-encoding pseudogene, was isolated by screening a human library. A sequence of 3004 bp, containing the GS coding region and both the 5' and 3' flanking sequences, was identified that exhibits all the characteristics of a processed pseudogene. The coding region shows 93% identity with the human GS cDNA (hGS) sequence and contains two frame-shifts and two termination codons. The coding sequence is flanked by a 9-bp AT repeat and a putative polyadenylation site, AATAAA, at the 3' end. Primer extension analysis and S1 nuclease mapping showed a transcription start point (tsp) 62 bp upstream from the start codon indicating a shorter untranslated region than hGS. Transfection of HeLa cells with cat constructs containing portions of the 5' flanking sequence showed the presence of a functional promoter/enhancer within 200 bp of the tsp, independent of its orientation.
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PMID:Detection of a functional promoter/enhancer in an intron-less human gene encoding a glutamine synthetase-like enzyme. 787 83

We have investigated the expression and extracellular release of active, recombinant Mycobacterium tuberculosis glutamine synthetase (EC 6.3.1.2), an enzyme that is a potentially important determinant of M. tuberculosis infection and whose extracellular release is correlated with pathogenicity. The M. tuberculosis glutamine synthetase gene encodes a polypeptide of 478 amino acids; 12 such subunits comprise the active enzyme. Northern blot, nuclease S1, and primer extension analyses revealed glutamine synthetase specific transcripts of approximately 1,550 and 1,650 nucleotides produced under low and high nitrogen conditions, respectively. Expression of recombinant M. tuberculosis glutamine synthetase in Escherichia coli YMC21E, a glutamine synthetase deletion mutant, led to transcomplementation of the mutant but not to release of active enzyme. Expression in Mycobacterium smegmatis 1-2c, from the gene's own promoter, resulted in the release of >95% of all recombinant enzyme. No hybrid molecules containing M. tuberculosis and M. smegmatis glutamine synthetase subunits were detected. Native and recombinant exported and intracellular glutamine synthetase molecules were indistinguishable from one another by mass, N-terminal amino acid sequence, antibody reactivity, and enzymatic activity. Since M. tuberculosis glutamine synthetase is similar to other, strictly intracellular, bacterial glutamine synthetases and the DNA sequence upstream of the structural gene does not encode a leader peptide, the information to target the protein for export must be contained in its amino acid sequence and/or conformation.
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PMID:Expression and efficient export of enzymatically active Mycobacterium tuberculosis glutamine synthetase in Mycobacterium smegmatis and evidence that the information for export is contained within the protein. 927 31

The glutamine synthetase II (GSII, encoded by glnII) activity detectable in crude extracts from Streptomyces coelicolor is low compared to the activity of glutamine synthetase I (GSI, encoded by glnA) and to that of GSII from S. viridochromogenes. We have identified and sequenced a 3.9-kb BglII-BamHI fragment carrying the glutamine synthetase II gene (glnII) from S. coelicolor. Besides glnII, this region contains four ORFs (orf1-orf4). While homologues of orf1 and orf2 were also found in the glnII region of the S. viridochromogenes chromosome, this was not the case for orf3 and orf4, which encode a putative hydrolase and a transcriptional regulator (Ptr) of the MarR family, respectively. High-resolution S1 nuclease mapping showed that the S. coelicolor glnII gene is expressed from two overlapping promoters. The first comprises a vegetative promoter sequence and the second contains sequence elements that are recognized by Esigma31. Similar promoter structures were found upstream of the S. viridochromogenes glnII gene. The involvement of ptr in glnII regulation was studied by gel retardation assays. Recombinant Ptr interacted with the upstream region of ptr, but not with the promoter region of glnII. A ptr gene replacement mutant (S. coelicolor IP) was also constructed. RT-PCR analysis of RNA from wild-type S. coelicolor and the IP mutant demonstrated that expression of orf3 depends on Ptr. Thus, the difference in gene organization between S. coelicolor and S. viridochromogenes is not responsible for the difference in GSII activity.
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PMID:Transcriptional analysis of the gene for glutamine synthetase II and two upstream genes in Streptomyces coelicolor A3(2). 1112 50

Streptomyces coelicolor has an unusually large arsenal of glutamine synthetase (GS) enzymes: a prokaryotic GSI-beta-subtype enzyme (encoded by glnA), three annotated glnA-like genes of the GSI-alpha-subtype and a eukaryote-like glutamine synthetase II (encoded by glnII). Under all tested conditions, GSI was found to represent the dominant glutamine synthetase activity. A significant heat-labile GSII activity, which is very low to undetectable in liquid-grown cultures, was only detected in morphologically differentiating S. coelicolor cultures. Analysis of glnA and glnII transcription by S1 nuclease mapping and egfp fusions revealed that, on nitrogen-limiting solid medium, glnII but not glnA expression is upregulated. An OmpR-like regulator protein, GlnR, has previously been implicated in transcriptional control of glnA expression. Gel retardation analysis revealed that GlnR is a DNA-binding protein, which interacts with the glnA promoter. It is not autoregulatory and does not bind to the upstream regions of the glnA-like genes of the alpha-subfamily, nor to the glnII promoter in vitro. A second GlnR target was identified upstream of the amtB gene, encoding a putative ammonium transporter. amtB forms an operon with glnK (encoding a PII protein) and glnD (encoding a putative PII nucleotidylyltransferase) shown by S1 nuclease protection analysis and reverse transcription-polymerase chain reaction (RT-PCR). An amtB and glnA promoter alignment revealed a putative GlnR operator structure. Downstream of glnII, a gene encoding for another OmpR-like regulator, GlnRII, was identified, with strong similarity to GlnR. Gel shifts with GlnRII showed that the promoters recognized by GlnR are also targets of GlnRII. However, GlnRII also interacted with the glnII upstream region. Only inactivation of glnR resulted in a glutamine auxotrophic phenotype, whereas the glnRII mutant can grow on minimal medium without glutamine.
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PMID:Two transcriptional regulators GlnR and GlnRII are involved in regulation of nitrogen metabolism in Streptomyces coelicolor A3(2). 1240 12

In plants, glutamine synthetase (GS) is the enzyme primarily responsible for the assimilation of ammonia into organic nitrogen. In Phaseolus vulgaris a number of isoenzymic forms of GS are found, each of which consists of eight subunits of mol. wt 41 000-45 000. The GS subunits of P. vulgaris have previously been shown to be encoded by a small multigene family and a partial cDNA clone for a nodule-specific GS subunit has been obtained. We report here the isolation and nucleotide sequencing of two essentially full-length GS cDNA clones (pR-1 and pR-2) from a root cDNA library and the deduced amino acid sequences of the corresponding GS subunits (355 amino acid residues each). The coding sequences of pR-1 and pR-2 are closely related (80% nucleotide homology, 88% amino acid homology), but their 5'- and 3'-untranslated regions have diverged almost completely. Both pR-1 and pR-2 are related to, but distinct from, the nodule GS clone, pcPvNGS-01 (or pN-1). Hybridization to genomic Southern blots showed that the three GS mRNAs are encoded by three seperate genes and indicated the existence of a fourth class of GS gene. An S1 nuclease protection assay demonstrated the presence of R-1 and R-2 mRNA in both roots and leaves and confirmed that expression of the N-1 gene is nodule-specific. Expression of the R-1 and R-2 genes in the roots did not change significantly during nodulation. However, only the R-1 gene is expressed in the nodules themselves, indicating that the R-2 gene is specifically repressed during nodule development.
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PMID:Primary structure and differential expression of glutamine synthetase genes in nodules, roots and leaves of Phaseolus vulgaris. 1645 87