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:3.1.27.5 (RNase)
17,967 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Undegraded rat liver polysomes were obtained after homogenizing the tissue in a medium containing NH4Cl, heparine, and yeast tRNA. Purification of poly(A)-containing RNA from polysomal RNA was accomplished by affinity chromatography on oligo(dT)-cellulose columns. Poly(A)-containing RNA molecules were monitored by the formation of ribonuclease-resistant hybrids with [3H]poly(U). To improve the separation of messenger RNA and ribosomal RNA by oligo(dT)-cellulose it was found essential to dissociate the aggregates formed between both molecular species by heat treatment in the presence of dimethylsulfoxide (Me2SO) prior to chromatography. Sucrose gradient analysis under denaturing conditions showed that the preparations obtained were virtually free of ribosomal RNA. Poly(A)-containing RNA constituted approx. 2.2% of the total polysomal RNA and the number average size was 1500--1800 nucleotides, as judged by sedimentation analysis on sucrose density gradients containing Me2SO. Approximately 8.2% of the purified preparation obtained was able to anneal with [3H]poly(U); the number average nucleotide length of the poly(A) segment of the RNA population was calculated to be 133 adenylate residues. Based on these values, our preparations appear to be greater than 90% pure. The RNA fractions obtained after oligo(dT)-cellulose chromatography were used to direct the synthesis of liver polypeptides in a heterologous cell-free system derived from wheat-germ. The system was optimized with respect to monovalent and divalent cations, and presence of polyamines (spermine). More than 65% of the translational activity present in the unfractionated polysomal RNA was recovered in the final poly(A)-containing RNA fraction. However, about 25% of the activity was found to be associated with the unbound fraction which was essentially free of poly(A)-containing RNA. Immunoprecipitation analysis with a specific antiserum to rat serum albumin demonstrated that about 6--8% of the labeled synthetic products translated from the poly(A)-containing RNA sample corresponded to serum albumin. Analysis of the translation products by sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed a heterogeneous distribution of molecular sizes ranging from 15 000 to greater than 70 000 daltons. Spermine not only increased the overall yield and extent of protein synthesis, but also resulted in higher yields of large protein products. Under optimal translation conditions a discrete peak representing about 7% of the total radioactivity was observed to migrate with rat serum albumin.
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PMID:Isolation and characterization of poly(adenylic acid)-containing messenger ribonucleic acid from rat liver polysomes. 66 61

A preparation of purified polyribosomes was isolated from wheat embryos germinating for 1 h with [3H]adenosine. Treatment of polyribosomes with pancreatic ribonuclease leads to the liberation of particles containing poly(A) sequences. Poly(A)-containing complexes show a heterogeneous distribution in the region 8--15 S. The ability of 8--15 S material to be absorbed to membrane filters under the conditions of low ionic strength and its sensitivity to pronase indicate that poly(A) sequences in polyribosomes are associated with protein. Analysis of this protein by electrophoresis in a polyacrylamide gel demonstrates the presence of two polypeptides with the molecular weights equal to 86,000 and 52,000.
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PMID:Proteins bound to poly(A) sequences of polyribosomes from germinating wheat embryos. 73 78

Precursor mRNA (pre-mRNA) was extracted from erythroid enriched bone marrow cells of the rabbit by the methods of Georgiev and Mantieva modified by Markov and Arion and of Holmes and Bonner, respectively. Density gradient centrifugation, base analysis and the effects of alpha-amanitin and actinomycin D on the synthesis of the cellular RNA showed signs of degradation in the rRNA-free 85 degrees C-fraction of the preparation according to Georgiev and Mantieva and a substantial rRNA contamination of the 65 degrees C-fraction. This RNA-fraction as well as the total RNA-preparation extracted according to Holmes and Bonner was purified from rRNA by affinity chromatography on poly(U)-Sepharose. Poly(A)+-RNA of all size-classes, among it a substantial amount of high molecular weight RNA (greater than 45 S), was isolated by this purification procedure. Especially the extraction according to Holmes and Bonner yields high molecular weight material but the critical step of this procedure often resulting in degradation of the RNA is the DNase treatment of the heavily DNA-contaminated total RNA-preparation either due to RNase contamination of the DNase or to the existence of RNase in the less intensive deproteinized RNA. The investigated cellular system is characterized by a very intensive rRNA synthesis which is typical for cells in the early stages of hematopoiesis. In contrast to investigations with purified RNA-polymerases and subcellular systems, but in accordance with data of in vivo experiments, alpha-amanitin inhibits both the pre-mRNA and the pre-rRNA synthesis.
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PMID:Pre-mRNA from erythroid enriched bone marrow cells of the rabbit. II. Characterization of pre-mRNA isolated by phenol extraction and poly(U)-Sepharose chromatography. 74 68

Pre-mRNA from bone marrow of rabbits enriched in erythroid cells was analyzed by T1 and pancreatic RNase treatment and poly(U)- and poly(A)-Sepharose chromatography to contain poly(A)-, oligo(U)- and double stranded sequences. The length of the poly(A)- and oligo(U)-sequences was determined by polyacrylamide gel electrophoresis using poly(A)- and oligo(U)-standards of defined length. Poly(A) from poly(A)+pre-mRNA isolated according to the method of Holmes and Bonner shows a size distribution between 40 and 130 nucleotides with an average of 75 nucleotides. Hot phenol extraction according to Georgiev et al. leads to a smaller size of about 25 nucleotides. The oligo(U)-segment consists of 80% U and is about 25 nucleotides long. Poly(A)+ pre-mRNA of about 12000--16000 nucleotides posseses 1--2 oligo(U)-units and one double strand of about 70 nucleotide pairs. Most (greater than 90%) of the oligo(U)-and the double stranded sequences are localized at least 1700 nucleotides away from the 3'terminus. Double strands were investigated with respect to their reannealing behaviour. The material consists of two types of double strands: 20% which reassociate at a cot/2 cot/2 of 1.3 . 10(-4) represent only one or a few types of double strands, the remaining 80% reassociate at a cot/2 of about 7 . 10(-2) and are more complex. Under hybridization conditions pre-mRNA molecules are able to self-annealation. 10% of the sequences become RNase stable.
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PMID:Pre-mRNA from erythroid enriched bone marrow cells of the rabbit. III. Poly(A)-, oligo(U)- and double stranded sequences. 74 69

Copolymers of vinyl bases with acrylic acid and styrene or 1-vinyluracil with maleic acid were found to stimulate in vitro polyphenylalanine synthesis using a system extracted from Escherichia coli MRE600. Poly(styrene-maleic acid) was found to inhibit a ribosomal bound ribonuclease. Poly(1-vinyluracil, maleic acid), poly(1-vinyluracil, acrylic acid), and poly(9-vinyladenine, acrylic acid) were not inhibitors of the ribosome bound ribonuclease. The potent (up to fivefold) stimulation by these three polymers is due to the action of the polymers to interfere with ribosomal bound inhibitory protein. A protein, removed by washing ribosomes with 1 M ammonium chloride, characterized by M.J. Miller, A. Niveleau, and A.J. Wahba ((1974) J. Biol. Chem. 249, 3803) has been described as a potent inhibitor of in vitro poly(U)-coded protein synthesis using extracts of Escherichia coli MRE 600.
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PMID:Mechanims of stimulation of in vitro protein synthesis by some copolymers of styrene, vinyluracil, and vinyladenine with maleic acid and acrylic acid. 78 18

Poly(A)-containing mRNAs labeled with [methyl-3H]methionine were isolated from nucleated erythroid cells obtained from the spleens of anemic mice. The RNAs were further separated into non-globin poly(A)-containing RNAs and highly purified globin mRNA by globin cDNA-cellulose affinity chromatography. DEAE-Sephadex column chromatography of the T2 ribonuclease digestion products of the cDNA-purified globin mRNA fraction yielded methylated resistant fragments with charges of -4.7 (Cap 1) and -5.3 (Cap 2). Digestion of the non-globin RNA fraction revealed a similar pattern with the addition of a methylated mononucleotide identified as 6-methyladenosine at -2 charges. Alkaline phosphatase treatment of the T2 resistant fragments reduced their charges by approximately 2, which is consistent with the removal of one terminal phosphate. Treatment of the globin T2 and alkaline phosphatase-resistant fragments withpenicillium P1 nuclease and alkaline phosphatase yielded a P1-resistant core structure in both fragments. In addition to the core, 2'-O-methylcytidine (Cm) was released from the more negatively charged globin fragment. The P1-resistant cores of the cap structures eluted from DEAE-Sephadex with the known standard m2G5'ppp5'Am and were found to be pyrophosphatase-sensitive establishing a 5'-5'-triphosphate linkage. The pyrophosphatase and alkaline phosphatase digestion products of the globin Cap 1 and Cap 2 core structures were analyzed by high voltage electrophoresis and paper chromatography and found to be 7-methyiguanosine (m7G) and the dimethylated nucleoside 6-methyl-2'-O-methyladenosine (N6mAm). A small amount of the singularly methylated adenosine, 2'-O-methyladenosine (Am) was also observed. The predominant sequences of the methylated nucleosides in the globin cap structures are therefore m7G5'ppp5'N6mAm and m7G5'ppp5'N6mAmpCm.
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PMID:Methylated nucleosides in globin mRNA from mouse nucleated erythroid cells. 83 41

Poly(xanthylic acid) [X)n] abolished the ability of (A)n-(U)n to induce interferon in "superinduced" (actinomycin D and cycloheximide) primary rabbit kidney cells. Under the same conditions, (X)n had a relatively minor effect on the interferon inducing capacity of (I)n-(C)n. Evidence based on mixing curves, melting profiles, pancreatic ribonuclease resistance and sucrose gradient ultracentrifugation pointed to the conclusion that the following three reactions occur depending on stoichiometry: (1) 2(A)n-(U)n + (X)n leads to (A)n-2(U)n + (A)n-(X)n; (2) (A)n-(U)n + (X)n leads to (A)n-(U)n-(X)n; (3) (A)n-(U)n + 2(X)n + (X)n-(U)n. The second reaction represents the formation of a new triple helix which can also be formed according to the following reactions: (X)n-(U)n + (A)n leads to (A)n-(X)n-(U)n; (A)n-(X)n + (U)n leads to (A)n-(X)n-(U)n.
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PMID:Polyadenylic-polyxanthylic-polyuridylic acid triple helix. 84 4

Several procedures were used to disassemble rat liver rough microsomes (RM) into ribosomal subunits, mRNA, and ribosome-stripped membrane vesicles in order to examine the nature of the association between the mRNA of bound polysomes and the microsomal membranes. The fate of the mRNA molecules after ribosome release was determined by measuring the amount of pulse-labeled microsomal RNA in each fraction which was retained by oligo-dT cellulose or by measuring the poly A content by hybridization to radioactive poly U. It was found that ribosomal subunits and mRNA were simultaneously released from the microsomal membranes when the ribosomes were detached by: (a) treatment with puromycin in a high salt medium containing Mg++, (b) resuspension in a high salt medium lacking Mg++, and (c) chelation of Mg++ by EDTA or pyrophosphate. Poly A-containing mRNA fragments were extensively released from RM subjected to a mild treatment with pancreatic RNase in a medium of low ionic strength. This indicates that the 3' end of the mRNA is exposed on the outer microsomal surface and is not directly bound to the membranes. Poly A segments of bound mRNA were also accessible to [(3)H] poly U for in situ hybridization in glutaraldehyde-fixed RM. Rats were treated with drugs which inhibit translation after formation of the first peptide bonds or interfere with the initiation of protein synthesis. After these treatments inactive monomeric ribosomes, as well as ribosomes bearing mRNA, remained associated with their binding sites in microsomes prepared in media of low ionic strength. However, because there were no linkages provided by nascent chains, ribosomes, and mRNA, molecules were released from the microsomal membranes without the need of puromycin, by treatment with a high salt buffer containing Mg++. Thus, both in vivo and in vitro observations are consistent with a model in which mRNA does not contribute significantly to the maintenance of the interaction between bound polysomes and endoplasmic reticulum membranes in rat liver hepatocytes.
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PMID:Release of poly A(+) messenger RNA from rat liver rough microsomes upon disassembly of bound polysomes. 88 10

Urine contains nondialyzable inhibitors of calcium oxalate crystal growth. We have pursued the hypothesis that these inhibitors may, in part, be acidic peptides and polyribonucleotide fragments. Homopolyribonucleotides and RNA inhibit calcium oxalate crystal growth at 5 x 10(-6) M of constituent ribonucleotide, whereas the monomer nucleotides are inactive at 10(-4) M. Poly-L-aspartic or glutamic acid are also inhibitory at 5 X 10(-6) M of amino acid, whereas the monomeric amino acids are inert. Gastric pepsin, a naturally occurring acidic peptide, is inhibitory. Incubation with nonspecific protease reduced the inhibitory effectiveness of normal human urine consistently and significantly, a fact compatible with an important contribution of peptides. A variable additional reduction was produced by subsequent treatment with ribonuclease, suggesting only a small role for polyribonucleotide. Sequential ion exchange and gel filtration chromatography and preparative disc gel electrophoresis yielded inhibitory material enriched with peptides that were strongly acidic and high in proline. Peptides and ribonucleotides seem to contribute to urinary nondialyzable crystal growth inhibitory activity.
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PMID:Acidic peptide and polyribonucleotide crystal growth inhibitors in human urine. 92 Aug 14

[3H]Poly(U) hybridizes very rapidly to polytene DNA from Drosophila hydei. When hybridization is performed at 30 degrees C in 2 X SSC to a large excess of DNA, 95% of the poly(U) becomes ribonuclease resistant. Also, complementary RNA transcribed in vitro from polytene DNA hybridizes to poly(U). 023--0.25% of the DNA is composed of (dA)-rich sequences and 0.23--0.31% of cRNA hybridizes to [3H]poly(U). The length of the (dA)-rich sequences on the DNA and cRNA is 40 nucleotides. The Tm values of these hybrids formed between DNA or cRNA-poly(U) is 45 degrees C. The poly(A) fragments from cytoplasmic RNA ranged from 80 to 170 nucleotides in lenght, and migrated in polyacrilamide gels as a broad peak. The average sizes of the poly(A) fragments from the poly(A)-containing RNA transcribed by nuclei isolated from salivary glands in vivo or in vitro were 40, 70, 170 and 70 nucleotides, respectively. Hybridization in situ of [3H]-poly(U) to chromosome squashes indicated that the (dA)-rich sequences are randomly distributed over the whole genome.
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PMID:Size and distribution of polyadenylic acid sequences in Drosophila polytene DNA and RNA. 92 97


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