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 carbamoylphosphate synthetase-aspartate transcarbamylase-dihydroorotase (CAD) gene encodes a tri-functional protein catalyzing the first three steps in de novo pyrimidine biosynthesis. Studies correlating CAD gene expression with cellular proliferation indicate the importance of understanding the regulation of the CAD gene. As a first step, the structure of the promoter region of the Syrian hamster CAD gene has been determined. Sequence analysis of 1671 base pairs of DNA revealed that the CAD promoter region is very GC rich. Primer extension analysis indicated that the transcription initiation site of the CAD gene is downstream from two GC boxes (consensus binding sites for the transcription factor Sp1). There is no TATA box appropriately spaced upstream from the transcription initiation site. Using RNase protection mapping, S1 nuclease analysis, and comparison to consensus splice donor/acceptor sites, the 5' end of the CAD gene has been determined to consist of a 241-base pair first exon, a 187-base pair first intron, a 140-base pair second exon, and a second intron that extends at least three kilobase pairs. Using conditions optimized for this GC-rich promoter, accurate transcription can be achieved in vitro. Analysis of CAD promoter deletions indicated that sequences extending only 114 base pairs upstream and 225 base pairs downstream from the transcription initiation site are sufficient for accurate and efficient transcription in vitro. DNase I footprinting reactions using this promoter fragment have identified three regions that bind proteins in a HeLa nuclear extract.
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PMID:Characterization of the 5' end of the growth-regulated Syrian hamster CAD gene. 198 61

The attenuation model for transcriptional regulation of the Escherichia coli pyrBI operon is based on the assumption that transcription terminates upstream of the structural genes at a rho-independent terminator when cells contain high levels of UTP. When, however, the cells are limited for pyrimidines, the presence of ribosomes translating the short leader peptide is presumed to cause an alteration in the secondary structure of the terminator in a way that allows RNA polymerase to transcribe the entire operon. These two premises of transcriptional regulation were tested by using exonuclease protection assays to map the 3' ends of transcripts extracted from cells containing either ample or depleted concentrations of pyrimidines. The results support the model since 99% of the pyrBI transcripts terminated at the (G + C)-rich region of dyad symmetry upstream of the structural genes when cells were grown in excess uracil. In addition, a significant portion (36%) of the pyrBI transcripts extracted from cells containing reduced pyrimidine concentrations extended past the dyad into the structural genes. This observation correlated with the amounts of aspartate transcarbamoylase synthesized in cells under the various conditions. The mapping technique was also used to determine the position of the 5' ends of the transcripts to measure contributions of two potential start sites (P1 and P2) to the pool of pyrBI transcripts. The results show that under all conditions no more than 3% of the total transcripts had 5' ends corresponding to the upstream promoter, P1. In cells lacking P1 virtually all transcripts from P2 terminated at the (G + C)-rich hairpin when the cellular level of pyrimidines was high. Conversely 57% of the transcripts extended past the terminator when cells were grown in UMP. The S1 nuclease technique also provided a measure of the steady state level of transcripts originating at P2. In cells depleted of pyrimidines there was a 5-10-fold increase in these transcripts depending on the number of copies of pyrBI. This increase, which is independent of attenuation, is caused by a different regulatory mechanism which as yet has not been identified.
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PMID:Attenuation in the regulation of the pyrBI operon in Escherichia coli. In vivo studies of transcriptional termination. 267 Sep 23

The first committed reaction in pyrimidine biosynthesis in Escherichia coli and Salmonella typhimurium is catalyzed by the allosteric enzyme aspartate transcarbamoylase (aspartate carbamoyltransferase; carbamoylphosphate:L-aspartate carbamoyltransferase, EC 2.1.3.2), the product of the pyrB-pyrI operon. Regulation of the pyrimidine pathway is achieved in part by changes in the enzyme's catalytic activity as a function of the concentration of substrates and other metabolites as well as by variations in enzyme synthesis in response to changes in cellular levels of pyrimidine nucleotides. Although there is substantial evidence that UTP concentration has a marked influence on expression of the pyrB-pyrI operon, the mechanism of this control is not known. We have cloned the operon and determined the nucleotide sequence of the region preceding the first structural gene (pyrB). These studies show two regions sharing considerable homology with the consensus sequence of E. coli promoters, a segment that can code for a 44-amino-acid leader peptide, and a sequence very similar to that of the attenuator of the trp operon. RNA transcripts from several bacterial strains were studied by S1 nuclease mapping. Under conditions leading to extensive enzyme synthesis there was a large production of transcript whose 5' end correlated with the putative promoter closer to the structural genes. At low levels of operon expression there was little transcript in the extracts and both promoters appeared to serve as initiation sites. The results are interpreted in terms of transcriptional control of the pyrB-pyrI operon according to an attenuation model that differs in novel ways from the mechanisms proposed for the regulation of amino acid biosynthesis.
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PMID:Synthesis of aspartate transcarbamoylase in Escherichia coli: transcriptional regulation of the pyrB-pyrI operon. 629 85