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:3.1.30.2 (endonuclease)
18,621 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The nucleoprotein organization of herpes simplex virus type 1 (HSV-1) DNA during productive infection was analyzed using staphylococcal nuclease. Both prior to and during the genome replication phase of infection, digestion of nuclei revealed two readily discernible forms of viral DNA, resistant and sensitive. The identity of these forms was established by the use of a variety of assays, including velocity sedimentation, nucleic acid hybridization and restriction endonuclease digestion and by employing temperature sensitive (ts) mutants impaired in either DNA replication or encapsidation of progeny DNA. Thus, nuclease resistant DNA was derived from encapsidated unit length genomes while sensitive DNA represented digestion products of replicating viral genomes. Importantly, no evidence was obtained for the arrangement of either parental or progeny viral DNA in nucleosomes. These findings are discussed with regard to the nucleoprotein structure of replicating viral DNA.
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PMID:Intracellular organization of herpes simplex virus type 1 DNA assayed by staphylococcal nuclease sensitivity. 216 12

Binding mechanisms of ADPR-transferase to restricted double-stranded DNA fragments of SV40 and pBR322 DNA were determined by nuclease protection techniques. Top and bottom strands of double-stranded DNA were identified by specific labeling with 32P. Protection against specific exonucleases identified binding of ADPR-transferase to DNA termini, whereas binding to internal regions of linear DNAs was probed by protection against endonucleases. ADPR-transferase protein protected against exonucleolytic attack from lambda exo and exoIII in all DNA fragments tested, demonstrating that the enzyme protein binds indiscriminately to all DNA termini. Extending earlier results [Sastry, S.S., & Kun, E. (1988) J. Biol. Chem. 263, 1505-1512], the modifying effect of the binding of ADPR-transferase to DNA induced changes in DNA conformation, as evident from altered pause sites that appeared following digestion of DNA fragments by lambda exonuclease in the presence of ADPR-transferase. In contrast to the nonselective binding of ADPR-transferase to DNA termini, ADPR-transferase conferred protection endonuclease attack (DNase I and micrococcal nuclease) only to the 209-bp EcoRI-PstI SV40 DNA fragment. These results indicate that binding of ADPR-transferase to relatively rare internal regions of restricted DNA fragments exhibits some degree of specificity. Specificity of binding appears to be related to the coincidental relative A+T-rich regions in DNA, and to DNA bending, both identified in the 209-bp SV40 DNA fragment. Synthetic polydeoxyribonucleotides containing dA-dT bind ADPR-transferase stronger than polydeoxyribonucleotides containing dG-dC. It was deduced from endonuclease protection patterns that binding of the enzyme protein leaves no defined footprints on the 209-bp SV40 DNA fragment, but there is significant modification of DNA structure following binding of the enzyme protein. Methylation protection assays and the prevention of the binding of ADPR-transferase to T4 DNA by its glucosylation indicate that the enzyme binds in the major groove of DNA. The 36-kDa A peptide fragment of ADPR-transferase [Buki, K. G., & Kun, E. (1988) Biochemistry 27, 5990-5995] exhibits the same protection against endonucleolytic enzymes as the intact ADPR-transferase molecule.
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PMID:Binding of adenosine diphosphoribosyltransferase to the termini and internal regions of linear DNAs. 250 40

Chromatin fragmentation of bovine peripheral blood lymphocytes from normal animals and the ones suffering from chronic lympholeucosis (CLL) by DNase I, micrococcal nuclease and purified Ca/Mg-dependent endonuclease from nuclei of human splenocytes was studied. The lymphocytes chromatin from CLL animals was shown to be more resistant to nucleases, than the one from normal animals. It was found that difference between fragmentation of chromatin samples from normal and CLL bovines was more dramatic when Ca/Mg- dependent endonuclease was used versus traditionally exploited DNase I and micrococcal nuclease. The data suggest that purified Ca/Mg-dependent endonuclease can be a useful enzymatic probe for detection of lymphocytes chromatin changes during CLL.
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PMID:[Ca/Mg-dependent endonuclease as a probe for detecting chromatin changes in lymphocytes in chronic lymphoid leukemia]. 282 38

Evidence is presented that endonuclease digestion of isolated, unfixed chromosomes results in the production of banding patterns similar to those produced by digestion of fixed, air-dried chromosomes. Mouse L cell chromosomes were isolated under acidic or relatively neutral pH conditions, exposed in situ (as wet mounts on glass slides) or in vitro (in suspension) to micrococcal nuclease, Alu I or Eco RI, treated with a buffered salt solution, and stained with Giemsa. After any of these endonuclease treatments in situ, the centromeric regions of the chromosomes were intensely stained, characteristic of the C-banding observed in fixed chromosomes exposed to the same treatments. Although the fixed chromosomes were morphologically well-preserved after endonuclease digestion, the morphology of chromosomes digested in situ was variable, ranging from normal to swollen to highly distorted chromosomes. In the latter, the endonucleases induced dispersion of non-C-band chromatin; however, C-bands were still apparent as condensed, differentially-stained regions. Exposure of isolated chromosomes to Alu I in vitro also resulted in well-defined C-banding and led to the extraction of about 70% of the chromosomal DNA. From these results, the mechanism of endonuclease-induced C-banding appears to involve the dispersion and extraction of digested chromatin.
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PMID:Endonuclease banding of isolated mammalian metaphase chromosomes. 301 70

Described here is an assay permitting exclusive quantitation of the kinetics of endonucleolysis by a mixed-function nuclease at substrate DNA concentrations much lower than those necessary with other assay methods. It makes possible determination of Michaelis parameters Km and kcat for endonuclease activity of such enzymes. The sensitivity of the assay method to very low DNA concentrations is obtained through use of 32P-labeled DNA as substrate. Digested DNA is electrophoresed, and computerized analysis of an autoradiogram made from the gel gives the extent of digestion. The analysis produces a profile of weight fraction vs. DNA fragment size for each sample taken from a nuclease-DNA reaction mixture. Each experimental profile is compared to a family of theoretical profiles generated by computer using theory of polymer statistics and assuming random cleavage. Theoretical profiles are found whose shapes most closely match those of the experimental profiles. Associated with each best-fitting theoretical profile is a value of the number of cuts made in the DNA sample. These values, plotted against time, give initial reaction velocities. Initial velocities from experiments at different DNA concentrations have been used to obtain Km and kcat for micrococcal nuclease under specific reaction conditions.
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PMID:Sensitive quantitation of endonuclease kinetics. 301 78

We have identified an endonuclease(s) that preferentially cleaves the internucleosomal linker regions in the aleurone chromatin producing mono- and oligonucleosomes. This enzyme(s) has been designated as a "linker"-specific nuclease(s). This nuclease does not require divalent cations for activity, and therefore it is not the "Ca2+-Mg2+-DNase" found in mammalian cells. The linker-specific nuclease activity is not detectable in the dry aleurone tissue and in the tissue treated with 0.5 mM cordycepin. The endonuclease activity of the aleurone tissue incubated with gibberellic acid is higher than the level of this endonuclease in tissue treated with abscisic acid or water alone. Nuclei isolated from embryos have lower levels of endonuclease activities compared to those from aleurone tissue. Digestion of the nuclei from embryos with micrococcal nuclease revealed the subunit structure of chromatin. In Southern blots of the HindIII digests of DNA from embryos, five DNA bands hybridized to a nick-translated alpha-amylase cDNA clone. In similar autoradiograms with aleurone DNA, particular bands are less visible, notably in the DNA isolated from the tissue treated with gibberellic acid. This is the first report of the presence of a linker-specific nuclease activity in plant cells.
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PMID:Study of barley endonucleases and alpha-amylase genes. 301 70

A gentle chromatin fractionation procedure was used to investigate solubility properties of Drosophila hsp70 heat-shock genes. After a brief digestion of isolated nuclei with micrococcal nuclease, most DNA is readily solubilized under low-ionic-strength conditions that maintain native nucleosomal organization. Actively transcribing hsp70 genes, however, are found to be enriched in the insoluble nuclear residue. Inactive genes are not resistant to solubilization, showing a fractionation pattern similar to that of bulk DNA. The insolubility characteristic correlates well with two other structural features of active hsp70 chromatin: increased sensitivity to endonuclease attack and disruption of the nucleosomal repeat pattern. The 5'-flanking regulatory region of active hsp70 genes is particularly resistant to solubilization, suggesting a role for binding of transcription factors in mediating this effect.
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PMID:Selective insolubility of active hsp70 gene chromatin. 308 25

To study the mechanism of gene repression by DNA methylation, M13 gene constructs were methylated to completion and inserted into mouse L cells by DNA-mediated gene transfer. All unmethylated sequences, regardless of their source, integrated into the DNA in a potentially active DNAase I-sensitive conformation. Total CpG methylation prevented the formation of this structure and rendered these sequences DNAase I-insensitive over the entire methylated domain. Whereas unmethylated DNA demonstrated additional conformational features of active genes, such as DNAase I hypersensitivity and restriction endonuclease-sensitive segments, these markers were not present when methylated DNA was used for transfection. The use of micrococcal nuclease to probe for active or inactive supranucleosome particles also showed that DNA methylation directs DNA into an inactive type of structure. The results suggest that DNA methylation may exert its effect on gene transcription by altering both specific and nonspecific interactions between DNA and nuclear proteins.
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PMID:DNA methylation affects the formation of active chromatin. 345 76

With the use of a reconstituted poly(ADP-ribosyl)ating enzyme system and three purified nucleases, micrococcal nuclease (MN), bull seminal RNase (BS RNase) and Ca2+, Mg2+-dependent endonuclease (BS DNase), as model acceptor proteins for ADP-ribose, the effect of ionic strength on the modification reaction was examined in detail. When these three nucleases were extensively poly(ADP-ribosyl)ated in this system at a low ionic strength (5 mM Tris), they were all inhibited by about 80% and the chain length of the polymer covalently bound to the nucleases was 13 to 23 ADP-ribose units. The observed inhibition was markedly prevented by increasing the ionic strength in the reaction mixture with a concomitant decrease in the polymer size bound to the nucleases. The NaCl concentrations required for decreasing the extent of the inhibition to half of the maximum were calculated to be 20, 50, and 100 mM for MN, BS RNase, and BS DNase, respectively. These values are similar to the NaCl concentrations required for decreasing the average chain lengths of the polymer to half, suggesting that the length of polymer is closely correlated to the extent of inhibition of these nucleases. DNA-binding affinities of these nucleases, expressed in terms of the NaCl concentrations required for eluting the enzymes from DNA-cellulose, were 140, 280, and 340 mM for MN, BS RNase, and BS DNase, respectively. Considering that maintainance of a ternary complex of poly(ADP-ribose) synthetase, acceptor and DNA may be essential for the modification reaction, the relatively strong salt effect observed in the modification of MN may be explained by its low DNA-binding affinity.
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PMID:Effect of ionic strength on chain elongation in ADP-ribosylation of various nucleases. 371 Oct 53

Conformational changes in the chromatin of the cerebral hemisphere of 3-, 14- and 30-day old developing rats were studied before and after its ADP-ribosylation using DNase I and micrococcal nuclease (MNase). The rate and extent of digestion of chromatin by DNase I are the highest at 3-day and decline progressively thereafter. The rate and extent of digestion by MNase do not change during development. ADP-ribosylation of chromosomal proteins was carried out by incubating nuclei with NAD+ for 30 min and was followed by endonuclease digestion. Both the rate and extent of digestion by DNase I and MNase were enhanced after ADP-ribosylation which was the maximum for 3-day rats.
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PMID:ADP-ribosylation induced changes in the conformation of the chromatin of the brain of developing rats. 396 86


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