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)

We have investigated some characteristics of the sarcoplasmic reticulum (Ca2+ + Mg2+)-dependent ATPase (Ca2+-ATPase) mRNA from smooth muscle using specific cDNA probes isolated from a rat heart cDNA library. RNA blot analysis has shown that the Ca2+-ATPase mRNA expressed in smooth muscle is identical in size to the cardiac mRNA but differs from that of fast skeletal muscle. S1 nuclease mapping has moreover shown that the cardiac and smooth muscle isoforms possess different 3'-end sequences. These results indicate that a distinct sarcoplasmic reticulum Ca2+-ATPase mRNA is present in smooth muscle.
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PMID:(Ca2+ + Mg2+)-dependent ATPase mRNA from smooth muscle sarcoplasmic reticulum differs from that in cardiac and fast skeletal muscles. 283 Oct 89

An extract capable of selectively transcribing early vaccinia virus genes was prepared by disrupting purified vaccinia virions and passing the soluble material through a DEAE-cellulose column to remove endogenous DNA. Runoff transcripts of predicted size were synthesized by using double-stranded DNA templates that contained truncated early vaccinia virus genes, whereas several late vaccinia virus genes were not transcribed under these conditions. Proper dilution of the enzyme extract was critical, and a threshold concentration of DNA was required. At 30 degrees C, runoff transcripts were detected after 5 min and synthesis slowed appreciably after 30 min. Mg2+ was the preferred divalent cation, and KCl concentrations above 20 mM were inhibitory. Correct initiation of transcription was demonstrated by high-resolution analysis of S1 nuclease-digested hybrids formed by annealing in vitro-synthesized RNA with 5'-end-labeled DNA. A requirement for a 31-base-pair transcriptional regulatory sequence was found by using templates with deletions in an early promoter region. This in vitro system may be useful for mapping early transcriptional initiation sites, measuring the effects of additional promoter mutations, and isolating transcription factors.
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PMID:Transcription of vaccinia virus early genes by a template-dependent soluble extract of purified virions. 299 63

In the absence of DNA aggregation, spermidine inhibited the relaxation of negatively supercoiled DNA by Escherichia coli topoisomerase I at concentrations of the polyamine normally found intracellularly. Spermidine also curtailed the cleavage of negatively supercoiled ColE1 DNA by the enzyme in the absence of Mg2+. On the contrary, knotting of M13 single-stranded DNA circles catalyzed by topoisomerase I was stimulated by the polyamine. Relaxation of supercoiled DNA by eukaryotic type 1 topoisomerases, such as calf thymus topoisomerase I and wheat germ topoisomerase, was significantly stimulated by spermidine in the same range of concentrations that inhibited the prokaryotic enzyme. In reactions catalyzed by S1 nuclease, the polyamine enhanced the digestion of single-stranded DNA and inhibited the nicking of negatively supercoiled DNA. These results suggest that spermidine modifies the supercoiled duplex substrate in these reactions by modulating the degree of single strandedness.
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PMID:Differential modulation by spermidine of reactions catalyzed by type 1 prokaryotic and eukaryotic topoisomerases. 300 Apr 18

RNA-dependent RNA polymerase activities were detected in purified particles of white clover cryptic viruses 1 and 2. The polymerases of the two viruses had different requirements for optimum activity. Enzyme activity was dependent upon the presence of virus particles, Mg2+, and the four ribonucleoside triphosphates, and was insensitive to actinomycin D, alpha-amanitin, and rifampicin. The labeled reaction products were dsRNAs as indicated by CF 11 column chromatography and by their ionic-strength-dependent sensitivity to hydrolysis by RNase A and resistance to S1 nuclease. The dsRNAs synthesized in vitro had the same electrophoretic mobilities as the corresponding viral templates.
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PMID:RNA-dependent RNA polymerase activity in two morphologically different white clover cryptic viruses. 335 1

Partially purified carnation cryptic virus (CarCV) preparations possessed RNA-dependent RNA polymerase activity which was absent in comparable preparations from virus-free carnations. Enzyme activity was dependent upon the presence of virus particles, Mg2+, and the four ribonucleoside triphosphates, and was insensitive to inhibitors of DNA-dependent RNA polymerases. The 32P-labeled enzyme reaction products were largely dsRNAs as indicated by resistance to S1 nuclease and RNase A at high but not low ionic strength. The in vitro synthesized dsRNAs hybridized specifically with CarCV genomic dsRNAs, and the radioactive products present in the polymerase reaction mixture sedimented with the virus particles in sucrose density gradients. The data suggest that the RNA-dependent RNA polymerase associated with CarCV particles is a replicase which catalyzes the synthesis of copies of the genomic dsRNAs.
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PMID:In vitro synthesis of double-stranded RNA by carnation cryptic virus-associated RNA-dependent RNA polymerase. 338 65

We describe the use of an enzymic probe of RNA structure, T2 ribonuclease, to detect alterations of RNA conformation induced by changes in Mg2+ ion concentration and pH. T2 RNase is shown to possess single-strand specificity similar to S1 nuclease. In contrast to S1 nuclease, T2 RNase does not require divalent cations for activity. We have used this enzyme to investigate the role of Mg2+ ions in the stabilization of RNA conformation. We find that, at neutral pH, drastic reduction of the available divalent metal ions results in a decrease in the ability of T2 RNase to cleave the anticodon loop of tRNAPhe. This change accompanies an increase in the cleavage of the molecule in the T psi C and in the dihydrouracil loops. Similar treatment of Tetrahymena thermophila 5S ribosomal RNA shows that changes in magnesium ion concentration does not have a pronounced effect on the cleavage pattern produced by T2 RNase. T2 RNase activity has a broader pH range than S1 nuclease and can be used to study pH induced conformational shifts in RNA structure. We find that upon lowering the pH from 7.0 to 4.5, nucleotide D16 in the dihydrouracil loop of tRNAPhe becomes highly sensitive to T2 RNase hydrolysis. This change accompanies a decrease in the relative sensitivity of the anticodon loop to the enzyme. The role of metal ion and proton concentrations in maintenance of the functional conformation of tRNAPhe is discussed.
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PMID:RNA structure analysis using T2 ribonuclease: detection of pH and metal ion induced conformational changes in yeast tRNAPhe. 620 83

Negatively superhelical pNS1 DNA with a molecular weight of 2.55 MDa (4 kbp) was found to contain 13 specific, unbasepaired sites that are sensitive to a single-strand-specific S1 nuclease cleavage. The S1-cleavage occurred once at these sites. In the absence of added Mg2+, the topoisomerase I purified from Haemophilus gallinarum formed a complex with the superhelical pNS1 DNA which has a hidden strand cleavage. Extensive proteinase K digestion of the complex led to cleavage of the DNA chain. Then the proteinase K-cleaved product was digested with S1, which can cut the opposite strand at the preexisting strand cleavage to generate unit-length linear DNA. Restriction endonuclease analysis of the linear DNA shows that the topoisomerase-induced cleavage occurred once at ten specific sites on the DNA. The topoisomerase caused mainly single-strand cleavage at these sites, but infrequently also caused double-strand cleavage at the same sites. Of interest is the fact that these sites considerably coincide with the S1-cleavable, unbasepaired sites.
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PMID:Correlation of enzyme-induced cleavage sites on negatively superhelical DNA between prokaryotic topoisomerase I and S1 nuclease. 630 25

Histones isolated from several sources, either singly or in combination promote the renaturation of complementary single strands of DNA, as measured by the acquisition of resistance to S1 nuclease. The reaction is rapid (T1/2 less than 1 min), and is stoichiometric rather than catalytic. Renaturation is stimulated by Mg2+, Mn2+, and Ca2+, but is strongly inhibited by Zn2+. Crude extracts of early embryos of Drosophila melanogaster possess renaturation activity which is protease sensitive, heat-stable, and acid-soluble, suggesting that most or all of it can be attributed to histones. This observation thus provides a functional assay for histones that should prove useful in studies of chromatin and histone-DNA interactions, as well as for the identification and isolation of histones and histone-like proteins in crude extracts.
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PMID:Renaturation of DNA: a novel reaction of histones. 678 53

Multivalent cations condense DNA in vitro, but it had been thought that a valence of at least + 3 was required in aqueous solution. We have found that Mn2+ can produce toroidal condensates of supercoiled plasmid DNA, but not of linearized plasmid. Mg2+ does not cause condensation, and neither MgCl2 nor NaCl can negate the effect of MnCl2, indicating that the condensation mechanism with Mn is not primarily electrostatic. Supercoiled MnDNA is more extensively digested than the linear form by S1 nuclease. Supercoiling appears to cooperate with Mn2+ in stabilizing helix distortions and also provides a "pressure" that enhances lateral association.
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PMID:Condensation of supercoiled DNA induced by MnCl2. 781 99

Structural differences between native (modified) and in vitro transcribed (unmodified) Escherichia coli tRNA(Val) were explored by comparing their temperature-absorbance profiles as a function of magnesium ion concentration and by probing their solution conformation with single- and double-strand-specific endonucleases. In vitro transcribed tRNA(Val) has a less ordered structure as monitored by thermal melting profiles; its Tm is appreciably lower than that of native tRNA(Val) at all Mg2+ concentrations. Structure probing experiments with nuclease S1 and ribonuclease V1 show that the unmodified tRNA(Val) transcript is more susceptible to nuclease attack at low Mg2+ concentrations, particularly in the D- and T-loops, indicative of at least a partial disruption of D-loop/T-loop interactions. These experiments also provide evidence for temperature-dependent alternative conformations of the anticodon loop of native tRNA(Val). Modified nucleosides are essential for the stability of these conformers; they cannot be detected in the unmodified in vitro transcript. The observations suggest that post-transcriptional modifications in tRNA allow the adoption of unique conformations and act to stabilize those that are biologically active.
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PMID:Probing structural differences between native and in vitro transcribed Escherichia coli valine transfer RNA: evidence for stable base modification-dependent conformers. 817 44


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