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:3.1.27.1 (RNase)
16,360 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Bacteriophage phi 6 contains three segments of double-stranded RNA within a nucleocapsid. Plasmids containing cDNA copies of the large genomic segment direct the synthesis of viral proteins that assemble into procapsids in Escherichia coli or Pseudomonas phaseolicola. These structures are dodecahedral assemblages of proteins P1, P2, P4, and P7. We report in this paper that these particles are capable of packaging viral single-stranded plus-sense RNA in vitro. The packaging reaction requires the presence of ATP or dATP. Synthesis of minus strands takes place within this filled procapsid in the presence of all four nucleoside triphosphates. Packaged ssRNA is found to be protected from added ribonuclease.
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PMID:In vitro packaging of the bacteriophage phi 6 ssRNA genomic precursors. 201 38

The first ATP-dependent complex formed in pre-mRNA splicing is the prespliceosome, a 30 S complex. This reaction was investigated using partially purified fractions isolated from nuclear extracts of HeLa cells. Previous studies (Furneaux, H. M., Perkins, K. K., Freyer, G. A., Arenas, J., and Hurwitz, J. (1985) Proc. Natl. Acad. Sci. U. S. A. 82, 4351-4355) have shown that DEAE-cellulose chromatography of nuclear extracts yielded two fractions (fractions I and II, eluted at 0.2 and 1 M NaCl, respectively) which carried out pre-mRNA splicing only when combined. Fraction II, alone and in the presence of ATP, supported the formation of the 30 S complex. In this report, we have separated fraction II into ribonucleoprotein and protein-rich fractions by isopycnic banding in CsCl. The combination of these two fractions completely replaced fraction II in prespliceosome formation; when supplemented with fraction Ib (1 M NaCl Biorex fraction derived from fraction I), the preparations supported spliceosome formation; when supplemented with fraction I, they yielded spliced products. The CsCl fractions, like fraction II, efficiently converted pre-mRNA to the 30 S complex with high yields (30-70%). The 30 S complex was shown to contain pre-mRNA complexed to U2 small ribonucleoproteins and small amounts of U1 small ribonucleoproteins. The 30 S complex protected a 50-nucleotide region at the 3'-end of the intron from T1 RNase attack. This region included sequences spanning the branch site, the polypyrimidine stretch and the AG dinucleotide of the 3'-splice site. When the 30 S complex was first generated with partially purified fractions, followed by the addition of a large amount of poly(U) or unlabeled pre-mRNA, the 30 S complex could be chased into a 55 S spliceosome complex by the addition of fraction Ib. These results support the conclusion, initially derived from kinetic data, that the 30 S complex is a precursor of the 55 S complex.
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PMID:Assemblage of the prespliceosome complex with separated fractions isolated from HeLa cells. 213 50

To determine whether tRNA or aminoacyl-tRNA synthetase is responsible for spermine stimulation of rat liver Ile-tRNA formation, homologous and heterologous Ile-tRNA formations were carried out with Escherichia coli and rat liver tRNA(Ile) and their respective purified Ile-tRNA synthetases. Spermine stimulation was observed only when tRNA from the rat liver was used. Spermine bound to rat liver tRNA(Ile) but not to the purified aminoacyl-tRNA synthetase complex. Kinetic analysis of Ile-tRNA formation revealed that spermine increased the Vmax and Km values for rat liver tRNA(Ile). The Km value for ATP and isoleucine did not change significantly in the presence of spermine. Furthermore, higher concentrations of rat liver tRNA(Ile) tended to inhibit Ile-tRNA formation if spermine was absent. Spermine restored isoleucine-dependent PPi-ATP exchange in the presence of rat liver tRNA(Ile), an inhibitor of this exchange. The nucleotide sequence of rat liver tRNA(Ile) was determined and compared with that of E. coli tRNA(Ile). Differences in nucleotide sequences of the two tRNAs(Ile) were observed mainly in the acceptor and anticodon stems. Limited ribonuclease V1 digestion of the 3'-32P-labeled rat liver tRNA(Ile) showed that both the anticodon and acceptor stems were structurally changed by spermine, and that the structural change by spermine was different from that by Mg2+. The influence of spermine on the ribonuclease V1 digestion of E. coli tRNA(Ile) was different from that of rat liver tRNA(Ile). The results suggest that the interaction of spermine with the acceptor and anticodon stems may be important for spermine stimulation of rat liver Ile-tRNA formation.
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PMID:Responsibility of tRNA(Ile) for spermine stimulation of rat liver Ile-tRNA formation. 233 46

Approximately one-third of the total ATP-hydrolysis activity in isolated HeLa nuclei is sensitive to RNAase (ribonuclease). This activity is selectively extracted with pulse-labelled RNA. In the extracts it co-sediments with various particles with sedimentation coefficients from 10S to 50S, but especially with 24S and 40S particles. ATP hydrolysis by the isolated particles was inhibited extensively (greater than 80%) by RNAase A, heparin and 0.2 M-NaCl. The activity of RNAase-treated particles was recovered when poly(A) was added, but not when DNA was added. The isolated particles exhibited RNAase-sensitive hydrolysis activities for dATP, GTP, CTP and UTP as well as for ATP, and the UTPase activity in the extracts showed nearly the same sedimentation distribution as the ATPase activity. When samples of isolated particles were irradiated with u.v. light in the presence of [alpha-32P]ATP, a 39 kDa polypeptide with a broad distribution from 10S to 50S like that of the ATPase and a 55 kDa polypeptide with a sharp distribution at 24S were photolabelled. Taken together, the data suggest that ATP-hydrolysis activity found in nuclear ribonucleoprotein subfractions appears to be the result of one or two RNA-dependent NTPases that are normally associated with endogenous RNA in a wide variety of particles.
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PMID:Characterization of a ribonuclease-sensitive nucleoside triphosphatase activity from HeLa nuclei. 240 2

More than 90% of rapidly-labelled nuclear RNA was associated with a nuclear matrix prepared from mouse leukemia L5178Y cells. The binding was not affected with up to 4 M NaCl; however, these RNAs were released from the nuclear matrix by treatment with a low ionic strength buffer (5 mM Tris-HCl buffer, pH 7.5, containing 1 mM ATP, 1 mM dithiothreitol, 0.2 mM ethylenediaminetetraacetic acid (EDTA) and 0.4 mM calcium chloride), without destruction of the sphere of the nuclear matrix. Actin filaments in the nuclear matrix were depolymerized with this buffer accompanied with rapidly-labelled RNAs. When the depolymerization was inhibited by slight modifications of the low ionic strength buffer (replacement of ATP by the same concentration of GTP; replacement of calcium ion by the same concentration of magnesium ion; addition of 20 micrograms/ml of phalloidine, which is a specific inhibitor of actin depolymerization), the release of rapidly-labelled RNAs from the nuclear matrix was also inhibited. The complex containing rapidly-labelled RNAs and matrix proteins was solubilized by a sonication from the nuclear matrix, and subjected to cesium chloride equilibrium centrifugation. Rapidly-labelled RNAs were concentrated on the bottom of the gradient accompanied with a small number of proteins (68K, 60K, 43K and 40K). The 43K protein was identified as actin by immunoblotting. By RNase digestion before equilibrium centrifugation, actin in the bottom fractions disappeared. These results suggest that rapidly-labelled RNAs anchor on the actin filaments in the nuclear matrix.
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PMID:Association of rapidly-labelled RNAs with actin in nuclear matrix from mouse L5178Y cells. 241 67

By crossing two strains of Saccharomyces cerevisiae deficient for each of the two methionine adenosyltransferase isoenzymes (ATP: L-methionine S-adenosyltransferase EC 2.5.1.6) respectively, we have constructed a strain strictly auxotrophic for S-adenosylmethionine and used it as a source of undermethylated mRNA suitable for in vitro transmethylation studies. RNA has been phenol-extracted from yeast cells shifted down to S-adenosylmethionine-free medium for 90 min and poly(A)-rich RNA has been prepared by oligo(dT)-cellulose chromatography. Upon incubation in vitro in the presence of methyl-labeled S-adenosylmethionine and mRNA (guanine-7-)-methyltransferase purified from wheat germ or yeast, undermethylated poly(A)-rich RNA became significantly labeled as compared to non-starved cells from the same strain, or from a wild-type control. Cap structures were resolved by paper chromatography afer T2 and P1 RNase digestion, and shown to be a mixture of m7G5'ppp5'G and m7G5'ppp5'A, irrespective of the enzyme source, in agreement with earlier in vivo studies in yeast mRNA capping and methylation.
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PMID:In vitro methylation of undermethylated yeast poly(A)-rich RNA using mRNA(guanine-7-)-methyltransferase purified from wheat germ or yeast. 241 1

A fragment of 16S RNA, cross-linked to S7 protein by UV irradiation of the 30S subunit of E. coli ribosome, was obtained by the action of T1 ribonuclease on the irradiated nucleoprotein. The digest was treated with polynucleotide kinase in the presence of [gamma-32P]ATP and the S7-cross-linked oligonucleotides were isolated. An individual oligonucleotide attached to S7 protein was obtained after proteinase treatment of the respective spot followed by electrophoresis. Sequencing of this oligonucleotide established its structure as 1233-1240 fragment of 16S RNA, the U1239 residue being the site of the S7 cross-linking. The developed general approach can be used for localizing protein - cross-linked residues in polynucleotides, whatever is the procedure employed for cross-linking.
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PMID:[General method of isolation and analysis of polynucleotide fragments cross-linked with proteins]. 241 22

The transformed androgen receptor from rat submandibular gland converts to a faster sedimenting form (6-8S) on a glycerol gradient centrifugation after withdrawal of a transformation-inducing reagent (KCl or ATP). In this report, the association of cytosolic RNA with the transformed androgen receptor was investigated as a possible mechanism of molecular conversion of the androgen receptor. When the transformed and converted androgen receptors were treated with RNase A, these receptors sedimented at 4.5S in a low-salt glycerol gradient. Addition of RNA from rat submandibular gland to the RNase-Sepharose-treated transformed receptor caused a shift of receptor peak from 4.5S to 5.8S. RNA from rat submandibular gland, yeast RNA and E. coli rRNA inhibited DNA-cellulose binding of a RNase-treated transformed receptor in the absence of molybdate. These observations suggest that conversion from the transformed 4S androgen receptor to a 6-8S form resulted from the association of RNA(s) with the transformed receptor.
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PMID:Ribonucleic acid association with androgen receptor from rat submandibular gland. 245 Feb 27

delta-Aminolevulinic acid is the first committed precursor in the biosynthesis of hemes, phycobilins, and chlorophylls. Plants and algae synthesize delta-aminolevulinic acid from glutamate via an RNA-dependent 5-carbon pathway. Previous reports demonstrated that cyanobacteria form delta-aminolevulinic acid from glutamate in vivo. We now report the direct measurement of this activity in vitro. Three oxygenic prokaryotes were examined, the unicellular cyanobacteria Synechocystis sp. PCC 6803 and Synechococcus sp. PCC 7002 (Agmenellum quadruplicatum PR-6) and the chlorophyll a- and b-containing filamentous prochlorophyte Prochlorothrix hollandica. delta-Aminolevulinic acid-forming activity was detected in soluble extracts of all three species. delta-Aminolevulinic acid formation by Synechocystis extracts was further characterized. Activity depended upon addition of reduced pyridine nucleotide, ATP, and Mg2+ to the incubation mixture. NADPH was a more effective pyridine nucleotide than NADH at low concentrations, but NADPH inhibited delta-amino-levulinic acid formation above 1 mM, whereas NADH did not. The pH optimum was about 7.6, and the ATP concentration optimum was 0.1 mM. Activity was stimulated by addition of RNA derived from Synechocystis or Chlorella, and abolished by preincubation with RNase A. After RNase inactivation, activity was restored by addition of RNasin to block further RNase action, followed by supplementation with Synechocystis RNA. Activity was inhibited by micromolar concentrations of hemin, as was previously found with plant and algal extracts. Complete dependence on added glutamate could not be achieved. Radioactivity was incorporated into delta-aminolevulinic acid when the incubation mixture contained 1-[14C]glutamate. Activity in the Synechocystis enzyme extract was stimulated by the addition of a partially purified enzyme fraction from Chlorella. It thus appears that prokaryotic oxygenic organisms share with chloroplasts the capacity for biosynthesis of photosynthetic pigments from glutamate via the RNA-dependent 5-carbon pathway.
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PMID:Transformation of glutamate to delta-aminolevulinic acid by soluble extracts of Synechocystis sp. PCC 6803 and other oxygenic prokaryotes. 245 30

We identified a Mg2+ dependent 5' exo-ribonuclease and an RNA ligase in cell-free extracts of Trypanosome brucei. The exo-ribonuclease in S100 or nuclear extracts, removes about 20 nts from the 5' end of SP6 derived capped as well as uncapped RNA and then stops. In contrast to the activity of the exo-ribonuclease on capped SP6 mini-exon transcripts, the exonuclease cannot degrade trypanosome-derived mini-exon transcripts or the mini-exon located at hsp 70 mRNAs. We therefore assume that the four secondary base modifications adjacent to the mini-exon cap, generated in vivo, confer resistance to the exo-ribonuclease. After exonuclease shortening of SP6 transcripts, an RNA ligase catalizes intramolecular ligation, generating a 3'-5' phosphodiester bond in a Mg2+ and ATP dependent reaction.
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PMID:A 5' exo-ribonuclease and RNA ligase of T. brucei. 246 Aug 26


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