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:3.1.30.1 (S1 nuclease)
3,660 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

An ovine mammary cDNA library has been constructed from total poly(A)+ RNA isolated from the mammary gland of a lactating ewe, using a classical procedure. Blunt-ended double-stranded cDNAs prepared with reverse transcriptase and nuclease S1 were tailed with dCTP, inserted into the dGMP-tailed PstI site of plasmid pBR322 and cloned in E. coli. Five series of homologous clones representing abundant messenger RNAs (strong hybridization with a single-stranded cDNA probe generated from total poly(A)+ RNA) were selected using each time a different predominant cloned ds-cDNA as probe, then identified by positive hybridization-translation of the cognate mRNA and subsequent immunoprecipitation and electrophoresis of the protein. The lengths of alpha s1-, alpha s2-, beta-, kappa-casein and beta-lactoglobulin mRNAs are in the range of 1.2, 1.1, 1.25, 1.0 and 0.85 kb, respectively, as determined by Northern blotting analysis. Five homologous mRNAs of similar sizes were identified in the porcine species by dot blot hybridization and Northern analyses. The nucleotide sequence of alpha s1-casein mRNA was determined by sequencing, according to Maxam and Gilbert, both a 1080 bp long cloned ds-cDNA and a ss-cDNA (268 nucleotides) generated by 5' extension of a 5' terminal truncated radiolabeled fragment (83 bp) of the relevant ds-cDNA, used as primer for reverse transcription. The 3' non coding region (431 nucleotides, excluding the poly(A) tail) represents 70% of the length of the coding region (618 nucleotides) flanked by a 61 nucleotide 5' region. Comparison of sequences of ovine and bovine, rat and guinea-pig alpha s1-casein mRNAs has revealed a greater homology in the 3' and especially 5' non coding regions. In the reading frame, the conserved regions are essentially those corresponding to the signal peptide and phosphopeptide domains. The derived 206 amino acid sequence of ovine pre-alpha s1-casein differs from that of its bovine counterpart (genetic variant B) by 24 amino acid substitutions and a deletion of 8 amino acid residues occurring in the polypeptide chain of the mature protein. Such a variation (84% homology only) in two phylogenetically closely related species indicates a high rate of evolution of alpha s1-casein.
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PMID:Construction and identification of recombinant plasmids carrying cDNAs coding for ovine alpha S1-, alpha S2-, beta-, kappa-casein and beta-lactoglobulin. Nucleotide sequence of alpha S1-casein cDNA. 300 1

The gene encoding the major capsid polypeptide (MCP 48) of frog virus 3 (FV 3) has been mapped on the viral DNA. Late FV 3 messenger RNA, hybrid-selected by the SalI-F fragment or a subset of these sequences, BamHI-L and -W fragments, directed the synthesis in vitro of a 48 000 mol. wt. (48K) polypeptide. This product was recognized by monospecific antibodies raised against the major capsid polypeptide. The RNA complementary to these DNA sequences was about 1350 nucleotides in size. This transcript, encoding MCP 48, was precisely located; S1 nuclease analysis indicated that its 5' end mapped at 1250 nucleotides to the right and its 3' end at 160 nucleotides to the left of the BamHI site at the junction between the BamHI-W and -L fragments.
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PMID:Mapping of the gene coding for the major late structural polypeptide on the frog virus 3 genome. 300 38

We have isolated and characterized a cDNA recombinant plasmid (pRLC429) specific for the rat heart myosin light chain 2 (MLC2). The cDNA insert consists of 446 base pairs, including a 72-base pair segment of the 3'-untranslated region. Additional 5'-sequence, not present in plasmid pRLC429, was obtained by primer extension of the cDNA. The extended cDNA sequence combined with the plasmid pRLC429 sequence provided the codon information for the entire MLC2 polypeptide and partial sequences for the 3'- and 5'-noncoding regions of MLC2 mRNA. The predicted amino acid sequence for rat heart MLC2 showed a high homology with the sequences available for the chicken (83%) and human heart (80%) MLC2s. However, the homology between rat heart MLC2 and its counterpart in rat skeletal muscle is relatively low (67%). On the basis of the nuclease S1 protection assay with uniformly labeled single-stranded pRLC429 DNA, subcloned into M13mp18 phage vector, we conclude that the rat atrial muscle also contains MLC2 of the ventricular type. In an attempt to ascertain whether structural variants of MLC2 are expressed in hypertrophic heart muscle, we examined the RNAs from spontaneously hypertensive rat where there is a natural progression of hypertrophy associated with an increase in blood pressure. The RNA isolated from 7-, 13-, and 18-week-old spontaneously hypertensive rat hearts protected the same length DNA against S1 nuclease as was observed with RNAs from the age-matched normal rat hearts, suggesting that there is a single MLC2 gene transcript expressed in both the normal and hypertrophic heart muscle cells.
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PMID:Heart myosin light chain 2 gene. Nucleotide sequence of full length cDNA and expression in normal and hypertensive rat. 300 70

Nucleases derived from Neurospora crassa mycelia with neutral single-strand (ss) endodeoxyribonuclease activity have been examined by immunochemical techniques and by sodium dodecyl sulfate - DNA gel electrophoresis. All of the intracellular nucleases, which have different divalent metal ion requirements, different strand specificities with single- and double-strand DNA, different modes of action on DNA and RNA, and other distinguishing characteristics, are immunochemically related to Neurospora endo-exonuclease. The evidence indicates that these enzymes are derived from one or more related large, inactive (precursor?) polypeptides that are first converted to 75- to 80-kdalton active polypeptide(s) which are very protease sensitive. Further limited proteolysis results in the production of the various active forms of nuclease studied here. Some proteolytic conversions may occur in a controlled manner in vivo in different cell compartments, but others are very likely artifacts resulting from uncontrolled proteolysis during extraction and isolation. The intracellular forms of Neurospora endo-exonuclease are immunologically cross-active with ss-DNA-binding nucleases isolated from Aspergillus nidulans and Saccharomyces cerevisiae. They are not immunochemically related to two extracellular Neurospora nucleases, the pancreatic DNase-I-like DNase A and a ss-specific exonuclease, and they are also not related to other fungal and plant nucleases with ss-specific endonuclease activity such as the S1 nuclease of Aspergillus oryzae, the P1 nuclease of Penicillium citrinum, and mung bean nuclease.
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PMID:An immunochemical study of Neurospora nucleases. 301 42

Polypeptide chain elongation factor 1 alpha (EF-1 alpha) of Saccharomyces cerevisiae is encoded by two distinct genes designated EF1 alpha A and EF1 alpha B [Nagata et al., EMBO J. 3 (1984) 1825-1830]. Both genes were cloned, and their nucleotide (nt) sequences were determined [see also Schirmaier and Philippsen, EMBO J. 3 (1984) 3311-3315, and Cottrelle et al., J. Biol. Chem. 260 (1985) 3090-3096]. They contain an open reading frame of 1374 nt coding for an identical protein of 458 amino acid residues, although their nt sequences differed at two positions. In this paper, we determined their 5'- and 3'-flanking sequences which were considerably different each other. From the S1 nuclease mapping of mRNA, both genes are found to be expressed almost to the same extent in exponentially growing cells. The transcription start points for EF1 alpha A and EF1 alpha B mRNAs were precisely located by primer extension procedure at 32 and 23 nt upstream of the start codons, respectively. The sequence which commonly exists in the 5'-flanking regions of ribosomal protein genes of S. cerevisiae was also present in the two EF1 alpha genes.
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PMID:Structure of the two genes coding for polypeptide chain elongation factor 1 alpha (EF-1 alpha) from Saccharomyces cerevisiae. 302 12

Herpes simplex viruses encode a structural protein which induces, in trans, expression of alpha genes, the first set of genes to be expressed after infection of permissive cells. This protein, designated as the alpha-trans-inducing factor (alpha-TIF), maps within the BamHI F fragment, and its gene has been sequenced. In the course of mapping the domain of the alpha-TIF gene, it was noted that the intact BamHI fragment was consistently more effective than the complete domain of the alpha-TIF gene in inducing expression of alpha genes. Cotransfections of DNA fragments containing an alpha indicator gene and the alpha-TIF gene with various regions of the BamHI F DNA fragment revealed that the sequences located 3' to the alpha-TIF gene raised the activity of the alpha-TIF gene to nearly the same level as that of the intact BamHI F fragment. The nucleotide sequence and S1 nuclease mapping analyses revealed the presence of two transcribed open reading frames capable of encoding polypeptides with translated molecular weights of 77,357 and 70,527. To determine whether the effect of these sequences in trans on alpha-TIF-mediated induction of alpha genes was due to expression of these genes or competition for transcriptional factors, we constructed plasmids that contained both genes. Into each or both of these genes we inserted, near the translation initiation sites, 14-base-pair linkers carrying translational stop codons (TAG) in all three reading frames. Analyses of these plasmids indicated that the gene encoding the 70,527-molecular-weight polypeptide reduced alpha-TIF-dependent induction of alpha genes, whereas the gene encoding the 77,357-molecular-weight polypeptide increased this activity. Insertion of the stop codons abolished these activities.
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PMID:Characterization and nucleotide sequence of two herpes simplex virus 1 genes whose products modulate alpha-trans-inducing factor-dependent activation of alpha genes. 302 33

A vaccinia virus (VV) gene required for DNA replication has been mapped to the left side of the 16-kilobase (kb) VV HindIII D DNA fragment by marker rescue of a DNA- temperature-sensitive mutant, ts17, using cloned fragments of the viral genome. The region of VV DNA containing the ts17 locus (3.6 kb) was sequenced. This nucleotide sequence contains one complete open reading frame (ORF) and two incomplete ORFs reading from left to right. Analysis of this region at early times revealed that transcription from the incomplete upstream ORF terminates coincidentally with the complete ORF encoding the ts17 gene product, which is directly downstream. The predicted proteins encoded by this region correlate well with polypeptides mapped by in vitro translation of hybrid-selected early mRNA. The nucleotide sequences of a 1.3-kb BglII fragment derived from ts17 and from two ts17 revertants were also determined, and the nature of the ts17 mutation was identified. S1 nuclease protection studies were carried out to determine the 5' and 3' ends of the transcripts and to examine the kinetics of expression of the ts17 gene during viral infection. The ts17 transcript is present at both early and late times postinfection, indicating that this gene is constitutively expressed. Surprisingly, the transcriptional start throughout infection occurs at the proposed late regulatory element TAA, which immediately precedes the putative initiation codon ATG. Although the biological activity of the ts17-encoded polypeptide was not identified, it was noted that in ts17-infected cells, expression of a nonlinked VV immediate-early gene (thymidine kinase) was deregulated at the nonpermissive temperature. This result may indicate that the ts17 gene product is functionally required at an early step of the VV replicative cycle.
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PMID:Nucleotide sequence and transcript organization of a region of the vaccinia virus genome which encodes a constitutively expressed gene required for DNA replication. 303 68

The nucleotide sequence of a 2-kilobase DNA fragment of the tdc region of Escherichia coli K-12, previously cloned in this laboratory, revealed two open reading frames, tdcC and ORFX, downstream from the tdcB gene (formerly designated tdc) encoding biodegradative threonine dehydratase. A 24-base-pair sequence separated tdcC from the dehydratase coding region, and an untranslated region of 60 nucleotides, which contains a recognizable -10 consensus sequence, was found between tdcC and ORFX. The deduced amino acid sequence of tdcC showed it to be a large hydrophobic polypeptide of 431 amino acid residues, whereas ORFX coded for a small 135-residue polypeptide lacking glutamine and tryptophan. A computer-assisted sequence analysis revealed no similarity among the tdcB, tdcC, and ORFX polypeptides, and a search of the GenBank database failed to detect similarity with any other known proteins. The tdc genes and ORFX showed similar codon usage and, in analogy with other bacterial genes, showed codon usage typical for genes expressed at an intermediate level. Transcriptional analysis with S1 nuclease indicated two distinct transcription start sites upstream of the tdcB gene in regions previously identified as promoterlike elements P1 and P2. Interestingly, expression of tdcB and tdcC, but not ORFX, was contingent upon the presence of P1. These results taken together tend to suggest that the biodegradative threonine dehydratase is the second gene in a polycistronic transcription unit constituting a novel operon (tdcABC) in E. coli implicated in anaerobic threonine metabolism.
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PMID:Molecular characterization of the tdc operon of Escherichia coli K-12. 305 59

The nucleotide sequence of an 878-bp BamHI-BglII restriction endonuclease fragment from citrate utilization transposon Tn3411 was determined, and was compared with that from plasmid pMS185 [Sasatsu et al., J. Bacteriol. 164 (1985) 983-993]. A long open reading frame for a 379-amino acid (aa) polypeptide (citB) was found 5' to the citA gene (431-aa membrane protein) in Tn3411 as well as in pMS185. Promoter regions were identified by RNA polymerase filter-binding assays, S1 nuclease mapping and cit-lac fusion experiments. The results indicated that two genes (citA and citB) have separate promoters, and the location of the promoter for the citB gene in the Tn3411 nucleotide sequence was different from that in pMS185. The regulation of transcription of the two genes (citA and citB) was characterized by the use of cit-lacZ fusions. The level of the citB promoter activity was about five-fold higher than that of the citA gene promoter, and transcription from both was not induced by citrate. Synthesis of the mRNA for the citB gene (especially with the wild-type Cit+ determinant) was suppressed by citrate, accompanying growth suppression of Escherichia coli. The citB gene expressed in E. coli minicells produced a membrane-associated 37.5-kDa polypeptide.
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PMID:Promoters and transcription of the plasmid-mediated citrate-utilization system in Escherichia coli. 306 41

We present a detailed analysis of the transcriptional products of the bithoraxoid (bxd) region of the Ultrabithorax domain in the bithorax complex of Drosophila melanogaster. This region is transcribed twice during development: between 3 and 6 hr of embryogenesis, a set of early transcripts, 1.1 to 1.3 kb in size, is synthesized; from the midthird larval instar through the adult stages, a late 0.8-kb transcript is synthesized. We have sequenced five cloned cDNAs representing early transcripts and three cDNAs representing the late transcript and have located their exons within the 40 kb of DNA comprising the bxd region. S1 nuclease protection and primer extension of both the early and late transcripts were used to further elucidate their structure. The early RNAs are produced by complex differential splicing of a series of exons derived from a 26-kb primary transcript. Curiously, these RNAs do not possess significant protein coding potential. The late bxd RNA comprises a single exon transcribed from an intronic region of the early transcription unit. This RNA, by contrast, possesses excellent coding potential and, if translated, would yield a 101-amino-acid polypeptide.
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PMID:Novel transcripts from the Ultrabithorax domain of the bithorax complex. 311 23


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