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)

1. Nuclei isolated from regenerating rat liver were digested with micrococcal nuclease and fractionated on glycerol gradients into soluble chromatin fragments and chromatin associated with the nuclear skeleton. 2. Distributions of DNA polymerases alpha and beta in these fractions were different. While beta polymerase followed closely the distribution of the chromatin fragments, alpha polymerase associated preferentially with the skeleton-chromatin complex. 3. At least 20% of total alpha polymerase in the nuclei was shown to be bound to the skeleton. In nuclei extracted with isotonic sucrose buffer containing 50 or 100 mM Tris-Cl the portion of the skeleton associated enzyme was increased to 40-50%. 4. These data show that the skeleton bound alpha polymerase was preferentially retained in the nuclei during salt extraction. 5. Contrary to the replicational DNA polymerase alpha, DNA polymerase beta did not show any affinity to the skeleton.
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PMID:Intranuclear localization of DNA polymerases alpha and beta in regenerating rat liver. 686 93

The physical properties of two types of androgen-binding sites in prostatic nuclei were compared and found to be identical. The first type was released from chromatin by micrococcal nuclease digestion and solution in 0.6 M NaCl; the second resisted such treatment and remained associated with nuclear structures. After in vivo administration of [1,2-3H]testosterone to 24-h castrated rats and sonication of purified nuclei, 90% of the nuclear radioactivity was extracted with nuclease/salt treatment and was found by sucrose density gradient analysis to be associated with a 3 S androgen receptor. If sonication was omitted, 50 to 60% of the nuclear radioactivity was recovered in the nuclease/salt-resistant pellets or bound to nuclear matrices. Mild digestion of either of these particulate fractions with trypsin resulted in the release of a 3 S androgen receptor. After in vitro isotope-exchange labeling with [1,2-3H]dihydrotestosterone, the sedimentation coefficient, steroid specificity, and dissociation constant of the androgen receptors released by trypsin digestion of nuclease/salt-resistant pellets or nuclear matrices were similar to those of the receptors extracted by nuclease/salt treatment. These results indicate first, that all androgen-binding sites in prostatic nuclei can be released, either with nuclease/salt or trypsin digestion procedures to yield a 3 S androgen receptor with uniform binding characteristics, and second, that the androgen receptors are distributed between two intra-nuclear pools--one containing about 10,000 molecules/nucleus sensitive to micrococcal nuclease digestion and salt and the other containing about 8,000 to 13,000 androgen receptors tightly bound to the nuclear matrix.
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PMID:Isolation of 3 S androgen receptors from salt-resistant fractions and nuclear matrices of prostatic nuclei after mild trypsin digestion. 686 57

A chromatin fraction enriched for Xenopus 5S RNA genes has been isolated by restriction endonuclease digestion and sucrose gradient velocity sedimentation. Soluble chromatin sedimenting at 70-80S contains approximately 50% of the oocyte-expressed 5S RNA genes and only 1.5-3% of total chromatin DNA; this represents a 15- to 30-fold purification of the 5S genes. Such chromatin isolated from somatic cells (blood and cultured kidney cells) retains the transcriptionally-inactive state of the oocyte-expressed 5S genes. Soluble chromatin from somatic cells prepared by micrococcal nuclease digestion also retains the inactive state of the oocyte-type 5S genes. It is likely that the level of chromatin structure responsible for inactivity of the oocyte genes in somatic cells is the nucleosome or short chains of nucleosomes and not supranucleosomal structures. The oocyte-type genes can be rendered transcriptionally active in somatic cell chromatin either by salt extraction of some chromosomal proteins or by treatment with the ion exchange resin Dowex A50W-X2. Alternatively, activation of these genes can be achieved by incubating somatic cell chromatin or nuclei with an extract prepared from Xenopus oocytes. This effect is not specific for 5S RNA genes as the transcription of other small RNAs (including pre-tRNA) is stimulated by the oocyte extract. The activating factor(s) is resistant to micrococcal nuclease, nondialyzable, heat labile and sensitive to trypsin; thus it is highly likely to be a protein or a group of proteins. Partial purification of the activating factor(s) has been achieved by ion exchange chromatography.
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PMID:Control of 5S RNA transcription in Xenopus somatic cell chromatin: activation with an oocyte extract. 686 64

An estrogen-responsive mouse Leydig cell tumor line (Tumor 124958) has been shown to contain only a low-affinity binder for estradiol in the cytosol fraction. This differed from the putative estrogen receptor in terms of its hormone-binding specificity as well as affinity. In addition, the possibility that an estrogen receptor-like molecule exists in the nuclei even without hormonal stimuli was examined using purified nuclei. Scatchard plot analyses showed that these nuclei possessed a large amount of estrogen binder having a high affinity for estradiol and diethylstilbestrol. The content of this nuclear binding component was not diminished by using molybdate, a potent inhibitor for receptor activation, and in vitro incubation of collagenase-dispersed cells with estradiol did not cause significant increase in the number of nuclear binding sites when compared with the values obtained by direct incubation of isolated nuclei with estradiol. These results support the view that this nuclear estrogen binder is not due to artificial migration of the cytosol receptor into nuclei during homogenization. The characterization of this nuclear binding component under cell-free conditions revealed that its affinity for estradiol in Mg2+-containing buffer was temperature dependent (Kd 3 nM at 30 degrees and 12 nM at 0 degrees) without significant alteration in the number of maximum binding sites. Introduction of a chelating agent (ethylenediaminetetraacetate) into the buffer system abolished the temperature effect on the affinity, resulting in high affinity for estradiol at both low and high temperatures. These Mg2+ and temperature effects were reversible. In addition, when compared with putative nuclear estrogen receptors, this nuclear binding was observed to be relatively resistant to high salt or micrococcal nuclease treatments in relation to solubilization from nuclei. However, trypsin digestion was found to result in a marked decrease in the nuclear binding sites, indicating that this unique nuclear binding component contains a protein unit(s). These results suggest the possibility that this tumor line contains a unique unoccupied nuclear estrogen binder which might be able to transmit estrogen signals to tumor cell nuclei with regard to tumor growth.
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PMID:Characterization of a unique nuclear estrogen-binding component in an estrogen-responsive mouse Leydig cell tumor. 687 50

The antigenicity and composition of chromatins differ markedly in chromatin preparations obtained by different procedures. Rat Novikoff hepatoma chromatin (NC) obtained by the "salt precipitation" and the micrococcal nuclease digestion procedures using significant levels of EDTA and NaCl each shows a common complement fixation (CF) capacity, exceeding chromatin preparations obtained from normal rat liver when tested with rabbit antisera raised to dehistonized NC. In contrast, "structured" NC preparations, which have been postulated to retain a native physical conformation, show minimal CF capacity when tested with the same antiserum but show high CF following elution of histones. While further progressive elution of non-histone proteins (NHPs) did not alter the CF capacity per unit DNA, the completely separated DNA and NHP fractions each showed minimal CF. The data suggest that the antigens detected in the CF assay predominantly represent an artifactual but specific complex of DNA and NHP arising from a denaturation of the native chromatin following elution of metal ions or histones. A qualitatively similar profile of NHPs in salt-precipitated NCs shows a range of total protein/DNA ratios, suggesting that the NHPs found in chromatin preparations may not be intrinsic to the native chromatin structure.
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PMID:Effects of divalent metal cations on composition and neoplasia-specific antigenicity of chromatins. 688 42

Chicken erythrocyte oligonucleosomes (trimers to about 20-mers) are able to interact with each other through the very lysine-rich histones (H1 and H5) and form heterogeneous globular particles with a mean diameter of about 300 A. These particles assemble spontaneously during micrococcal nuclease digestion of chromatin in the presence of 30 mM NaCl and contain approximately 25 nucleosomes. They are sensitive to ionic strength and unfold at lower salt concentrations but can be reconstituted by restoring the initial salt concentration. Even at 30 mM NaCl, the particles remain dynamic structures, being in equilibrium with their oligonucleosomal components as revealed by the fact that particle stability depends on the concentration of oligonucleosomes.
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PMID:Aggregation of small oligonucleosomal chains into 300-A globular particles. 693 58

A tandemly repeated DNA hexanucleotide sequence, 5'C-C-C-C-A-A3', that occurs at the termini of extrachromosomal DNA molecules coding for rRNA (rDNA) in Tetrahymena macronuclei was examined to determine whether it is packaged as nucleosomes. This repeated DNA sequence comprises the terminal few hundred base pairs at each end of the linear rDNA molecules. Digestion of macronuclei with micrococcal nuclease showed that this DNA sequence is protected from digestion but is left, following digestion, as a single but broad size class of DNA fragments several hundred base pairs long, under conditions in which bulk macronuclear DNA and rDNA were digested to fragments that were multiples of approximately 200 base pairs in length. The repeated C-C-C-C-A-A was found protected as fragments longer than the bulk macronuclear DNA digestion products at all times during digestion. Together with putative associated protein(s), this protected DNA was soluble after lysis of micrococcal nuclease-digested macronuclei at low salt concentrations but was insoluble in 0.075--0.2 M KCl, regardless of the extend of digestion. The size and solubility properties of the repeated C-C-C-C-A-A DNA nucleoprotein complex after micrococcal nuclease digestion of macronuclei are clearly distinguishable from those of nucleosomes, and it is inferred that this DNA sequence in macronuclei is packaged in chromatin by proteins other than histones.
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PMID:Non-nucleosomal packaging of a tandemly repeated DNA sequence at termini of extrachromosomal DNA coding for rRNA in Tetrahymena. 694 Dec 83

We have described earlier a chromatin-bound protease with unique specificity for histone H2A [Eickbush, T. H., Watson, D. K., & Moudrianakis, E. N. (1976) Cell (Cambridge, Mass.) 9, 785--792]. In the present study, we explore the nature of interactions that form and stabilize the enzyme-chromatin system by using the activity of the protease to monitor its binding to DNA and DNA-histone complexes. During salt extraction of chromatin, the protease is released at an ionic strength between that required for the extraction of the slightly lysine-rich histones (H2A and H2B) and the arginine-rich histones (H3 and H4). The reassociation of this nonhistone protein to DNA has an absolute requirement for the H3--H4 tetramer and is only enhanced by the H2A--H2B dimer in the presence of the tetramer. We believe that the binding of the enzyme onto DNA requires some histone-elicited compaction of the helix. We have also examined the distribution of this enzyme within the chromatin fiber by isolating pools of monomer nucleosomes from micrococcal nuclease digests of 0.6 M NaCl extracted chromatin and from reconstituted DNA-protein complexes. The H2A-protease is found with these monomer nucleosome pools, and no activity can be detected in the low molecular weight products released during the digestion. Thus, by virtue of its extraction characteristics from chromatin and its association with isolated nucleosomes, this nonhistone protein exhibits properties hitherto assigned only to the inner histones.
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PMID:Histone-dependent reconstitution and nucleosomal localization of a nonhistone chromosomal protein: the H2A-specific protease. 704 60

Chicken erythrocyte chromatin, whole or (H1,H5)-depleted, was dissociated in 2 M NaCl in the presence or absence of 5 M urea at pH 8 or pH 5, and reconstituted by decreasing the concentration of NaCl and urea. The reassociated products were characterized by their solubilities in 0.1 mM EDTA, micrococcal nuclease digestion, cross-linking of histones, and fractionation of histone oligomers by solubility in ammonium sulfate solution. In the absence of urea, the nucleosome structure and the histone octamer were reconstituted perfectly at both pH 5 and pH 8. When chromatin was exposed to urea, no nucleosome structure or histone octamer was obtained at pH 5 either decreasing the concentration of salt first or that of urea first. At pH 8, the chromatin structure was regained fairly well by decreasing the concentration of urea first, but only partially by decreasing the concentration of salt first. Solubility in 0.1 mM EDTA was found to be a good criterion for monitoring the proper reassociation of chromatin.
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PMID:Studies on histone oligomers. IV. Reassociation of chromatin from histones of various conformations. 707 56

Histone deposition in HeLa cells has been studied by monitoring the fractionation and electrophoresis mobility of pulse-labeled histones under conditions that separate newly replicated from bulk chromatin DNA. The separation efficiency of these two methods is approximately 70%. Following micrococcal nuclease digestion, chromatin was fractionated by salt elution. 50-65% of the newly synthesized histones eluted with bulk chromatin at NaCl concentrations between 0.1 and 0.3 M and were further down to co-electrophorese with bulk chromatin DNA, not with the more extensively digested newly replicated chromatin DNA contained in those fractions. The remaining chromatin fractions, solubilized with 0.4-0.6 M NaCl, were several-fold enriched in nascent DNA (Annunziato, A. T., Schindler, R. K., Thomas, C. A., Jr., and Seale, R. L. (1981) J. Biol. Chem. 256, 11880-11886) and were correspondingly enriched for the balance (35-50%) of newly synthesized core histones. This fraction of newly synthesized core histone may be preferentially deposited onto newly replicated DNA. In contrast, histone H1 showed little tendency toward deposition onto new DNA. Within 15 min all new core histones attained the same solubility and electrophoretic mobility as bulk chromatin. We conclude that newly synthesized histones are deposited onto both replicating and nonreplicating regions of chromatin.
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PMID:Association of newly synthesized histones with replicating and nonreplicating regions of chromatin. 708 80


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