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)

A cell free system that supports replication-dependent chromatin assembly has been used to determine the mechanism of histone deposition during DNA replication. CAF-I, a human cell nuclear factor, promotes chromatin assembly on replicating SV40 DNA in the presence of a crude cytosol replication extract. Biochemical fractionation of the cytosol extract has allowed separation of the chromatin assembly reaction into two steps. During the first step, CAF-I targets the deposition of newly synthesized histones H3 and H4 to the replicating DNA. This reaction is dependent upon and coupled with DNA replication, and utilizes the newly synthesized forms of histones H3 and H4, which unlike bulk histone found in chromatin, do not bind to DNA by themselves. The H3/H4-replicated DNA complex is a stable intermediate which exhibits a micrococcal nuclease resistant structure and can be isolated by sucrose gradient sedimentation. In the second step, this replicated precursor is converted to mature chromatin by the addition of histones H2A and H2B in a reaction that can occur after DNA replication. The requirement for CAF-I in at least the first step of the reaction suggests a level of cellular control for this fundamental process.
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PMID:Stepwise assembly of chromatin during DNA replication in vitro. 184 80

It is demonstrated that the histone (H3-H4)2 tetramer can find specific positions on DNA, even in the absence of other histones. Purified histone (H3-H4)2 tetramers were reconstituted onto 208-base-pair (bp) DNA molecules containing a nucleosome-positioning sequence by using salt-gradient dialysis. The stoichiometry of histone tetramer to DNA was shown to be 1:1. Digestion with micrococcal nuclease led to formation of protected DNA fragments of approximately 73 bp. Cleavage of the 73-bp DNA with restriction enzymes produced a small set of defined bands, demonstrating positioning of the (H3-H4)2 tetramer on DNA. Analysis of the restriction digests shows that the 73-bp DNA corresponds mainly to two fragments, one lying on either side of the pseudo-dyad axis of the major position adopted by complete histone octamers on this DNA. This result means that a single (H3-H4)2 histone tetramer can fold approximately 146 bp of DNA with the same positioning as the complete octamer but that a region near the pseudo-dyad is only weakly protected against micrococcal nuclease attack in the absence of histones H2A and H2B.
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PMID:Nucleosome positioning is determined by the (H3-H4)2 tetramer. 196 26

The preparation of hybrid histone octamers with wheat histone H2A variants replacing chicken H2A in the chicken octamer is described. The fidelity of the reconstituted hybrid octamers was confirmed by dimethyl suberimidate cross-linking. Polyglutamic-acid-mediated assembly of these octamers on long DNA and subsequent micrococcal nuclease (MNase) digestion demonstrated that, whereas chicken octamers protected 167 base-pairs (representing 2 full turns of DNA), hybrid histone octamers containing wheat histone H2A(1) with its 19 amino acid residue C-terminal extension protected an additional 16 base pairs of DNA against nuclease digestion. The protection observed by hybrid histone octamers containing wheat histone H2A(3) with both a 15 residue N-terminal and a 19 residue C-terminal extension was identical with that observed with H2A(1)-containing hybrid histone octamers with only the 19 residue C-terminal extension. These results suggest that the role of the C-terminal extension is to bind to DNA of the "linker" region. The thermal denaturation of chicken and hybrid core particles was identical in 10 mM-Tris.HCl.20 mM-NaCl, 0.1 mM-EDTA, confirming that there was no interaction between the basic C-terminal extension and DNA of the core particle. Denaturation in EDTA, however, showed that hybrid core particles had enhanced stability, suggesting that the known conformational change of core particles at very low ionic strength allows the C-terminal extension to bind to core particle DNA under these conditions. A model accounting for the observed MNase protection is presented.
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PMID:Extended C-terminal tail of wheat histone H2A interacts with DNA of the "linker" region. 202 50

Assembly of nucleosomes on relaxed, covalently closed DNA has been studied in a nuclear extract of Xenopus laevis oocytes. Nucleosomes containing the four histones H3, H4, H2A and H2B but lacking histone H1 are readily assembled on the DNA. The pattern of micrococcal nuclease digestion shows that the nucleosomes assembled in the absence of ATP and Mg (II) are closely packed, with a periodicity of 150 base pairs (bp). In contrast, in the presence of ATP and Mg (II) the spacing of the nucleosomes is 180 bp, similar to that observed for nucleosomes assembled on DNA microinjected into oocyte nuclei. The ATP and Mg (II) requirements for the assembly of correctly spaced nucleosomes are unrelated to the activity of the ATP and Mg (II) dependent DNA topoisomerase II in the extract; addition of specific inhibitors of eukaryotic DNA topoisomerase II has no effect on the spacing of the reconstituted nucleosomes. The ATP requirement in the assembly of correctly spaced nucleosomes can be substituted by adenosine 5'-O-3'-thiotriphosphate (gamma-S-ATP) but not by adenyl-5'-yl imidodiphosphate (AMP-P-(NH)-P).
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PMID:Assembly of correctly spaced chromatin in a nuclear extract from Xenopus laevis oocytes. 217 Sep 36

Mononucleosomes were released from both isolated mammalian (hog thyroid) and protozoan (Tetrahymena) nuclei by the bleomycin-induced DNA-strand breaking reaction. Trout sperm nuclei, on the other hand, were protected from the bleomycin-mediated DNA degradation. The mononucleosomes released from the bleomycin-treated nuclei contained the core histones H2A, H2B, H3, and H4; while HMG1 and HMG2 proteins, in addition to the core histones, were detected in the mononucleosomes obtained by micrococcal nuclease digestion of nuclei. HMGs, but not H1 histone, were dissociated into the supernatant by cleavage of chromatin DNA with bleomycin, whereas both HMGs and H1 were found in that fraction by digestion of nuclei with micrococcal nuclease. HMG1 and HMG2 were exclusively dissociated from chromatin with 1 mM bleomycin under the solvent condition where the DNA strand-breaking activity of the drug is repressed. These observations suggest the possibility that bleomycin preferentially binds to linker DNA regions not occupied by H1 histone in chromatin and exclusively dissociates HMG proteins and breaks the DNA strand. The results of the effects on bleomycin-induced DNA cleavage of nuclei of various drugs including polyamines, chelating agents, intercalating antibiotics such as mitomycin C or adriamycin, and radical scavengers are also presented.
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PMID:Release of nucleosomes from nuclei by bleomycin-induced DNA strand scission. 247 91

We have studied how non-histone proteins HMG1 and HMG2 interact with rat liver chromatin using reconstitution and chemical cross-linking procedures. Both proteins were found to associate to chromatin only to some extent and always with a marked preference for short oligonucleosomes, mainly mono- and dinucleosomes. However, a slight reconstitution with the long polynucleosomal fraction can be observed in H1-depleted chromatin. Reconstitution is non-random and a clear preference for regions highly sensitive to staphylococcal nuclease (EC 3.1.31.1) is observed. Chemical cross-linking has allowed us to identify H1, H2A and H2B as the histones contacted by HMG1 and HMG2 upon reconstitution. Also, we present evidence that HMG1 and HMG2 interact with the nucleosomal particle without replacing H1 or any other histone.
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PMID:Non-random reconstitution of HMG1 and HMG2 in chromatin. Determination of the histone contacts. 271 62

The Xenopus oocyte supernatant (oocyte S-150) forms chromatin in a reaction that is affected by temperature and by the concentration of ATP and Mg. Under optimal conditions at 27 degrees C, relaxed DNA plasmids are efficiently assembled into supercoiled minichromosomes with the endogenous histones H3, H4, H2A and H2B. This assembly reaction is a gradual process that takes four to six hours for completion. Micrococcal nuclease digestions of the chromatin assembled under these conditions generate an extended series of DNA fragments that are, on average, multiples of 180 base-pairs. We have examined the effect of histone H1 in this system. Exogenous histone H1, when added at a molar ratio of H1 to nucleosome of 1:1 to 5:1, causes an increase in the micrococcal nuclease resistance of the chromatin without causing chromatin aggregation under these experimental conditions. Furthermore, the periodically arranged nucleosomes display longer internucleosome distances, and the average length of the nucleosome repeat is a function of the amount of histone H1 added, when this histone is present at the onset of the assembly process. In contrast, no major change in the length of the nucleosome repeat is observed when histone H1 is added at the end of the chromatin assembly process. Protein analyses of the purified minichromosomes show that histone H1 is incorporated in the chromatin that is assembled in the S-150 supplemented with histone H1. The amount of histone H1 bound to chromatin is a function of the total amount of histone H1 added. We define here the parameters that generate histone H1-containing chromatin with native nucleosome repeats from 160 to 220 base-pairs, and we discuss the implications of these studies.
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PMID:Assembly and properties of chromatin containing histone H1. 281 Mar 66

The micrococcal nuclease cleavage sites have been mapped in the H2A coding and flanking regions of the sea-urchin histone DNA chromatin. A hypersensitive area, centered around - 100 base pairs from the H2A starting site, is found only in embryos actively transcribing the alpha-subtype histone genes. In mesenchyme blastula embryos, upon inactivation of the H2A gene, this region becomes protected while two other areas, near the transcription starting site and in the proximity of the 3' palindromic sequence, become preferential targets for the enzyme. Analysis of the pattern of micrococcal nuclease cleavage on the same region of the histone gene cluster in sperm and late blastula chromatin and on the corresponding segment of protein-free DNA indicates that distinct nucleosomal arrangements characterize the histone genes in the two cell populations.
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PMID:Different micrococcal nuclease cleavage patterns characterize transcriptionally active and inactive sea-urchin histone genes. 300 83

Bovine thymus and trout testis chromatin were fractionated into regions which differed in their micrococcal nuclease accessibility and solubility properties, and the distribution of the ubiquitinated histone species among these chromatin regions was elucidated. Ubiquitinated (u) species of histones H2A and H2B were enriched in the nuclease-sensitive, low-ionic-strength, soluble fraction of both chromatins. These results indicate that the presence of ubiquitinated histones may alter nucleosome-nucleosome interactions and destabilize higher-order chromatin structures. Bovine thymus chromatin was separated into aggregation-resistant, salt-soluble and aggregation-prone, salt-insoluble chromatin fractions. The aggregation-resistant chromatin fraction depleted in H1 histones was enriched in uH2A and uH2B, with uH2B showing the greater enrichment. The chromatin fragments were also stripped and reconstituted with the H1 histones prior to fractionation. The results were the same as above: uH2A and uH2B were preferentially localized in the aggregation-resistant. H1-depleted chromatin fraction, suggesting that chromatin regions enriched in ubiquitinated histone species have a reduced affinity for the H1 histones. Thus, ubiquitinated histone species may be one of the contributing factors in the differential assembly of various parts of the genome.
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PMID:The ubiquitinated histone species are enriched in histone H1-depleted chromatin regions. 304 Jan

The effects induced by Fe, Mn, or Mg deficiency or cold shock on the DNA content and histones of Euglena gracilis have been examined and compared to those produced by Zn deficiency. The DNA content of the stationary-phase organisms used as controls is 2.1 micrograms/10(6) cells. The DNA of stationary-phase iron-deficient (-Fe), magnesium-deficient (-Mg), manganese-deficient (-Mn), zinc-deficient (-Zn), and cold-shocked (CS) cells is increased to 3.0, 4.6, 6.2, 3.8, and 3.8 micrograms/10(6) cells, respectively. The electrophoretic mobilities of proteins solubilized with 0.4 N H2SO4 from CS, -Fe, -Mg, and -Mn cells are nearly identical and are characteristic of the five histone classes, H1, H2A, H2B, H3, and H4. In contrast, no histones are found in the equivalent acid extract from -Zn cells. The effect of micrococcal nuclease on chromatin from control, CS, and -Zn cells was examined. The chromatin of CS cells is 1.2-fold while that from -Zn cells is 10-30-fold more resistant to micrococcal nuclease digestion than is the chromatin of control cells. Thus, the chromatin of cells grown in Zn-deficient conditions differs markedly from that of organisms cultured in media deficient in Fe, Mn, or Mg or exposed to cold shock.
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PMID:Euglena gracilis chromatin: comparison of effects of zinc, iron, magnesium, or manganese deficiency and cold shock. 309 72


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