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)

Two approaches have been explored for the synthesis of double-stranded DNA from single-stranded DNA template complementary to rabbit 9S globin mRNA (cDNA). (i) cDNA was elongated with dCMP or dTMP homopolymeric tracts using terminal deoxynucleotidyltransferase (EC 2.7.7.31; nucleosidetriphosphate:DNA deoxynucleotidylexotransferase). cDNA-dC, in the presence of an oligo(dG)10 primer, was an efficient template with either DNA polymerase of Escherichia coli (EC 2.7.7.7; deoxynucleosidetriphosphate:DNA deoxynucleotidyltransferase) or RNA-directed DNA polymerase of avian myeloblastosis virus. cDNA-dT [ with an oligo(dA)10 primer] functioned as template only with E. coli polymerase. (ii) cDNA, without homopolymeric tails, was also efficiently copied in the absence of oligonucleotide primer, by DNA polymerase of avian myeloblastosis virus or of E. coli. The product of the reaction consisted of long hairpin molecules which could be converted into DNA duplex (melting temperature, 93 degrees) by digestion with single-strand nuclease S1. The data indicate that a loop structure on the 3' end of cDNA allowed DNA synthesis to take place by a "self-priming" mechanism. Some of the double-stranded DNA synthesized corresponded to the entire sequence of the 9S mRNA template. The synthesis of full-length double-stranded DNA from mouse globin mRNA and immunoglobulin light chain mRNA is also discussed.
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PMID:Stepwise biosynthesis in vitro of globin genes from globin mRNA by DNA polymerase of avian myeloblastosis virus. 6 60

The complete nucleotide sequence of a 1.8-kilobase DNA fragment containing the cell cycle-regulated thymidylate synthase gene (TMP 1) of the yeast Saccharomyces cerevisiae is presented. This analysis has revealed a 912-base pair open reading frame which encodes a 304-amino acid residue protein with a calculated Mr of 35,007. The tmp1-6 and cdc21-1 mutant alleles of this gene also have been sequenced, and both show single base pair changes which would result in different amino acid substitutions. The amino acid sequence of the yeast thymidylate synthase gene derived from the DNA sequence shows considerable homology when compared with the human, mouse, Herpesvirus saimiri, Leishmania major, Leishmania tropica, Escherichia coli, Lactobacillus casei, bacteriophage T4, and Bacillus subtilis phage phi 3T enzymes. Northern blot hybridization reveals that the TMP 1 mRNA is a 1.15-kilobase polyadenylated transcript. A set of consensus yeast mRNA splice sequences appears within the open reading frame of TMP 1, but S1 nuclease protection experiments reveal that splicing of the mRNA does not occur. Disruption of the gene by the introduction of a large insertion did not produce any defect besides the expected dependence on dTMP for growth. Specifically, the viability of the mutants in the presence of dTMP indicates that the protein does not play a significant structural role in a complex of replication enzymes.
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PMID:Molecular characterization of the cell cycle-regulated thymidylate synthase gene of Saccharomyces cerevisiae. 303 Oct 48

When the nonprotein chromophore of neocarzinostatin was allowed to react with either calf thymus DNA or poly(dA-dT) . poly(dA-dT) in the presence of 2-mercaptoethanol and the DNA was precipitated with ethanol, 5% of the fluorescence attributable to the naphthalene rings of the chromophore coprecipitated with the DNA. Most of this fluorescence remained attached to DNA through successive reprecipitations, suggesting formation of covalent adducts between chromophore and DNA. Enzymatically digested poly(dA-dT) . poly(dA-dT)-chromophore adduct contained, in addition to deoxyadenosine and thymidine, several highly fluorescent hydrophobic products, separable by reverse-phase chromatography, all of which contained both adenine and thymine radiolabel, as well as chromophore radiolabel. One such product consistently had twice as much thymine as adenine, suggesting a structure chromophore-d(TpApT), in which the attached chromophore rendered both phosphodiester bonds refractory to endonuclease S1. This adduct fragment was completely hydrolyzed at pH 12, releasing adenine, 3'-dTMP, and 5'-dTMP. At pH 7, the adduct fragment slowly released chromophore and 3'-dTMP with parallel kinetics, leaving a modified d(ApT), which was cleaved by snake venom phosphodiesterase to yield 5'-dTMP and a modified deoxyadenosine. These hydrolysis patterns are unlike those of any previously characterized base or phosphotriester DNA adduct but rather indicate an altered deoxyadenosine sugar. The formation of adducts containing a modified deoxyribose suggests that deoxyribose may be the site of covalent chromophore attachment. Alteration of this same site, possibly the 5'-carbon of the sugar moiety, may account for the extreme lability of the phosphodiester bond.
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PMID:Covalent adducts of DNA and the nonprotein chromophore of neocarzinostatin contain a modified deoxyribose. 621 Sep 7

The priA gene of the basidiomycete Lentinus edodes possesses a pyrimidine (CT)-rich stretch (26 bp) that includes a short (6-bp) repeat, the elements of which form a mirror repeat at and near the transcriptional initiation sites. A DNA fragment that included this sequence was inserted into pBR322, and the resulting plasmids were introduced into Escherichia coli. Analysis of the susceptibility of these pBR322 derivatives to cleavage by S1 nuclease, following isolation from E. coli, indicated the formation of an open, S1-sensitive structure within and just downstream of the CT/AG-biased sequence. Replacement of two dTMP residues in one of the repeat elements by dGMP resulted in the elimination of the S1-cleavable open structure from the plasmids. To analyze the effect of the CT/AG-biased sequence from priA in the basidiomycete Coprinus cinereus, the integrating vectors pLC2 and pLC2mutCT were used; these contained the wild-type priA promoter and the mutant priA promoter with the aforementioned mutation in the mirror repeat, respectively. The Streptomyces-derived bialaphos resistance gene (bar) was fused downstream of the promoters, and the resulting plasmids, pLC2-bar and pLC2mutCT-bar, were introduced into C. cinereus. Transformants carrying pLC2mutCT-bar grew significantly more slowly on bialaphos-containing agar plates and contained a noticeably lower level of the bar transcript when compared with the transformants obtained with pLC2-bar. These results suggest that an unusual structure induced by the CT/AG-biased sequence is required for efficient gene expression from the priA promoter.
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PMID:Structure and function of a pyrimidine/purine-biased sequence from the 5'-flanking region of the basidiomycete Lentinus edodes gene priA. 1077 44