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:3.1.31.1 (micrococcal nuclease)
2,818 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Bovine leukemia virus 60 to 70S RNA was heat denatured, the polyadenylic acid-containing species were separated by velocity sedimentation, and several size classes were translated in a micrococcal nuclease-treated cell-free system from rabbit reticulocytes. The major RNA species sedimented at 38S and migrated as a single component of molecular weight 2.95 x 10(6) when analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The predominant polypeptides of the in vitro translation of bovine leukemia virus 38S RNA were products with molecular weights of 70,000 and 45,000; minor components with molecular weights of 145,000 and 18,000 were also observed. Two lines of evidence indicate that the 70,000- and 45,000-molecular weight polypeptides represent translation products of the gag gene of the bovine leukemia virus genome (Pr70gag and Pr45gag). First, they are specifically precipitated by a monospecific antiserum to the major internal protein, p24, and second, they are synthesized and correctly processed into virion proteins p24, p15, and p10 in Xenopus laevis oocytes microinjected with bovine leukemia virus 38S RNA. The 145,000-molecular weight polypeptide was immunoprecipitated by the anti-p24 serum and not by an antiserum to the major envelope glycoprotein, gp60. It contained all the tryptic peptides of Pr70gag and additional peptides unique to it, and thus represents in elongation product of Pr70gag in an adjacent gene, presumably the pol gene. The 18,000-molecular weight product was antigenically unrelated to p24 and gp60 and shared no peptides in common with Pr70gag, Pr45gag, or the 145,000-molecular weight polypeptide. It was maximally synthesized on a polyadenylic acid-containing virion 16 to 18S RNA, and we present evidence that this RNA is a 3' end-derived subgenomic fragment of the bovine leukemia virus genome rather than a contaminating cellular RNA.
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PMID:Translation of bovine leukemia virus virion RNAs in heterologous protein-synthesizing systems. 8 18

We have recently shown that, after the histones and most of the nonhistone proteins are gently removed from HeLa metaphase chromosomes, the chromosomal DNA is still highly organized and relatively compact. The structure of these histone-depleted chromosomes is due to the presence of a number of nonhistone proteins that form a central scaffold that retains the approximate size and shape of intact chromosomes and to which the DNA is attached, predominantly forming loops. We now demonstrate that the protein scaffold may be isolated independently of the DNA by treating HeLa chromosomes with micrococcal nuclease before removing the histones.The chromosomal scaffolds may be isolated by sucrose density gradient centrifugation as a well-defined peak that is stable in 2 M sodium chloride, but is dissociated by treatment with proteases, 4 M urea, or 0.1% sodium dodecyl sulfate. Polyacrylamide gel electrophoresis reveals that the protein content of scaffold preparations is identical to that of histone-depleted chromosomes. Fluorescence microscopy of purified scaffolds in isolation buffer shows that the particles still possess the familiar chromosome morphology. When the scaffolds are examined in the electron microscope, a fibrous structure with the approximate size and shape of intact, paired chromatids is seen. Less than 0.1% of the chromosomal DNA and virtually no histones are associated with the purified scaffold structures.
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PMID:Isolation of a protein scaffold from mitotic HeLa cell chromosomes. 27 Jul 27

At the completion of mitosis, the two daughter cells are connected by a channel of membrane-bound cytoplasm, the intercellular bridge. This structure contains parallel arrays of spindle microtubules which are associated, at the bridge midline, with a metallophilic band called the midbody. In an effort to characterize midbody components, intercellular bridges were partially purified. The purification consisted of brief sonication of telophase populations of mouse L929 cells in order to shear intercellular bridges from daughter cells, digestion of chromatin by an excess of micrococcal nuclease, and differential centrifugation to enrich for intercellular bridges. Electron microscopy of these preparations substantiated the identity of the bulk of material as intercellular bridges. After solubilization with sodium dodecyl sulfate, the protein components of these preparations were iodinated with Na(125)I and separated by two-dimensional gel electrophoresis. From these analyses, the major polypeptide components of intercellular bridges appear to be tubulin, varying amounts of plasma membrane proteins, and a polypeptide with an apparent molecular weight of 42,000. Time-course studies reveal that this polypeptide is first detectable in a pelletable form approximately 30 min after cells are released from metaphase block, reaches maximal spot intensity in late telophase, and is no longer detectable in G1 populations. We interpret these data to suggest that the 42,000-dalton polypeptide is a component of the midbody.
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PMID:Partial purification and characterization of the intercellular bridge from cultured mouse cells. 28 7

The activity of ribonuclease P on precursor tRNA substrates from Escherichia coli can be abolished by pretreatment of this enzyme with micrococcal nuclease or pancreatic ribonuclease A, as well as by proteases and by thermal denaturation. Highly purified RNase P exhibits one prominent RNA and one prominent polypeptide component when examined in polyacrylamide gels containing sodium dodecyl sulfate. The buoyant density in CsCl of RNase P, 1.71 g/ml, is characteristic of a protein-RNA complex. The activity of RNase P is inhibited by various RNA molecules. The presence of a discrete RNA component in RNase P appears to be essential for enzymatic function. A model is described for enzyme-substrate recognition in which this RNA component plays an important role.
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PMID:Ribonuclease P: an enzyme with an essential RNA component. 35 97

Nuclei from hepatoma tissue culture (HTC) cells were isolated by standard methods and incubated in media commonly used for nuclease digestions (DNAase I and micrococcal nuclease) and for in vitro RNA synthesis. During the incubation, histones can be deacetylated from both control cells and cells treated with 6 mM sodium butyrate to enhance the levels of histone acetylation. Deacetylation of histone is much more apparent in nuclei isolated from sodium butyrate-treated cells. Inclusion of 6 mM sodium butyrate in the incubation medium effectively inhibits the endogenous deacetylase activity acting on histones H3 and H4, whereas sodium acetate at the same concentration has very little inhibitory effect.
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PMID:Histone deacetylation in nuclei isolated from hepatoma tissue culture cells. Inhibition by sodium butyrate. 42 71

Synthesis of phiX174 viral (+) strand circles in vitro requires gene A protein, rep protein, DNA binding protein, and DNA polymerase III holoenzyme (Eisenberg, S., Scott, J. F., and Kornberg, A., (1976) Proc. Natl. Acad. Sci. U.S.A. 73, 3151-3155). We have used this reaction as an assay to isolate gene A protein in greater than 90% purity. Its molecular weight under denaturing conditions is 59,000. The protein tends to aggregate and lose activity at low ionic strength. Tritium-labeled gene A protein cleaves the phiX174 duplex replicative form and is bound to it in a 1:1 ratio as part of an active replication complex. The attachment, at the 5' phosphoryl end of the cleavage point, is apparently covalent. The complex was not dissociated by: (i) banding in CsCl, (ii) treatment with 0.2 M NaOH, or (iii) boiling in 1% sodium dodecyl sulfate and electrophoresis on a sodium dodecyl sulfate-acrylamide gel; only micrococcal nuclease digestion of the DNA released the protein.
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PMID:Purification and characterization of phiX174 gene A protein. A multifunctional enzyme of duplex DNA replication. 44 53

Rat liver chromatin was digested by micrococcal nuclease. Chromatin subunits (or mononucleosomes) were isolated by sucrose density gradient and subsequently fractionated by 6% polyacrylamide gel electrophoresis into two major components. One component (MN1) of the mononucleosomes had a higher mobility, contained histones H2A, H2B, H3, H4, and shorter DNA fragments (140 base pairs) while the other (MN2) contained all five histones and longer DNA fragments (180 base pairs). Both submononucleosomes (MN1 and MN2) were found to contain nonhistone chromatin proteins (NHCP). By electrophoresis in 15% sodium dodecyl sulfate-polyacrylamide gel, 9 and 11 major fractions of NHCP were identified in the submononucleosomes MN1 and MN2, respectively. It was also observed that treatment of mononucleosomes with 0.6 M NaCl removes most of these NHCP and histone H1 except for two major NHCP which remain in the core particles.
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PMID:Identification of nonhistone chromatin proteins in chromatin subunits (or mononucleosomes) devoid of histone H1. 47 14

Buoyant-density centrifugation of unfixed chromatin has been performed in a newly devised medium containing 3-iodo-1,2-propanediol and metrizamide. Chromatins were obtained from isotopically labeled mouse hepatoma cells in suspension culture, either grown normally or density labeled in a medium containing bromodeoxyuridine, by mild digestion of isolated nuclei with micrococcal nuclease. When a mixture of normal and density labeled chromatin, marked with [14C]thymidine and [3H]bromodeoxyuridine, respectively, was centrifuged in the medium, chromatin peaks represented by labeled DNA were resolved to the extent expected from their separate banding profiles. Centrifugation of an equivalent chromatin mixture labeled with [14C] and [3H]lysine, respectively, also yielded resolution of chromatin peaks represented by labeled proteins. Only small amounts of labeled proteins were dissociated from chromatin in the gradient medium. Labeled proteins recovered from the gradient fractions were analyzed by electrophoresis on sodium dodecyl sulfate-polyacrylamide gels. The results suggested that most of the histones remained associated with the original stretches of DNA during the centrifual fractionation period. Essentially all of the dissociated proteins were found to be non-histone proteins.
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PMID:Fractionation of unfixed chromatin by buoyant-density centrifugation in gradients containing 3-iodo-1,2-propanediol and metrizamide. 64 Oct 28

I have grown HeLa cells in 5 mM sodium n-butyrate leading to extensive in vivo histone acetylation, and have characterized the structure of chromatin containing the modified histones. Nuclear DNA in butyrate-treated cells is digested 5-10 fold more rapidly by DNAase I than the DNA of control cells. Staphylococcal nuclease degrades the two nuclear samples to acid-soluble material with identical rates; this nuclease, however, does excise nucleosomes with extensively acetylated histones from the nucleoprotein chain preferentially. The physical properties of unsheared chromatin and isolated core particles from control and butyrate-treated cells are closely similar, as are the rates of digestion of core particles from the two cell preparations by DNAase I. Determination of the relative susceptibilities of cleavage sites for DNAase I demonstrates that the site 60 bases from the ends of the DNA resistant in control cells, becomes susceptible to the nuclease in core particles containing acetylated histones. Similarly, the 5' terminal phosphate at the end of DNA in core prticles is removed by staphylococcal nuclease 2-3 fold faster in particles containing acetylated histones than in particles from control cells.
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PMID:Structure of chromatin containing extensively acetylated H3 and H4. 65 72

Nuclei isolated from cultured Chinese hamster cells were treated with micrococcal nuclease and lysed, and the resulting chromatin subunit classes (nucleosomes) were purified by sedimentation and resedimentation through isokinetic sucrose gradients. Nucleosomes isolated from [3H]thymidine-labeled cells were analyzed for DNA size using both polyacrylamide gel and electron microscopic techniques. Nucleosomes isolated from [14C]lysine-labeled cells were analyzed for protein content using a sodium dodecyl sulfate-polyacrylamide gel system. The results from monitoring the [14c]lysine in each protein indicate that, in the nucleosome classes (monomer through tetramer), the molar ratios of histones H2A, H2B, H3, and H4 are equivalent. Furthermore, in each population of the nucleosome classes monomer through tetramer, it was possible to demonstrate that this histone unit (H2A + H2B + H3 + H4) is present, on the average, in the amount of two for monomers, four for dimers, six for trimers, and eight for tetramers. This is direct experimental confirmation of the prediction of R.D. Kornberg [(1974) Science 184, 868] concerning the substructure of chromatin.
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PMID:Histone composition of nucleosomes isolated from cultured Chinese hamster cells. 91 4


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