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)

We have compared the nucleosomal organization of c-Ha-rasVal 12 oncogene-transformed NIH-3T3 fibroblasts with that of normal fibroblasts by using micrococcal nuclease (MNase) as a probe for the chromatin structure. The bulk chromatin from asynchronously and exponentially growing ras-transformed cells was much more sensitive to MNase digestion than chromatin from the normal cells. Southern hybridization analyses of the MNase digests with probes specific for the ornithine decarboxylase (odc) and c-myc genes showed that the coding and/or 3' end regions of these growth-inducible genes carry a nucleosomal organization both in ras-transformed and normal cells. Studies with cells synchronized by serum starvation showed that in both cell lines the nucleosomal organization of chromatin is relatively condensed at the quiescent state, becomes highly decondensed during the late G1 phase of the cell cycle, and starts again to condense during the S phase. However, in ras-transformed cells the decondensation state stayed much longer than in normal cells. Moreover, irrespective of the phase of the cell cycle the bulk chromatin as well as that of the odc and c-myc genes was more sensitive to MNase digestion in the ras-transformed cell than in the normal fibroblast. Decondensation of the chromatin was also observed in the normal c-Ha-ras protooncogene-transfected cells, but to a lesser extent than in the mutant ras-transformed cells. Whether the increased degree of chromatin decondensation plays a regulatory role in the increased expression of many growth-related genes in the ras-transformed cells remains an interesting object of further study.
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PMID:c-Ha-rasVal 12 oncogene-transformed NIH-3T3 fibroblasts display more decondensed nucleosomal organization than normal fibroblasts. 219 41

The properties of a Saccharomyces cerevisiae 20 kDa polypeptide (Yp20) and its relationship to human ras antigen were tested. Yp20 was isolated from commercial yeast cells by the procedure of Sommer (1978). Proteins associated with yeast chromatin were released by micrococcal nuclease digestion and purified by sucrose gradient centrifugation. Rabbit polyclonal and mouse monoclonal antibodies specifically detecting the Yp20 antigen have been generated. We observed that Yp20 was recognized by anti-ras polyclonal and monoclonal antibodies. Mammalian Ha-ras and Ki-ras proteins were specifically detected by anti-Yp20 antibodies. Based on immunological cross-reactivities, we believe that Yp20 may share some homology with the yeast YP2 gene previously described. Anti-Yp20 antibodies will be used to isolate the gene that encodes the protein. Practical applications of our antibodies for the detection of tumor specific antigens will be discussed.
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PMID:Yeast as source of oncoproteins. 266 75

The chromatin structure of the human c-K-ras gene has been investigated in various cultured normal and tumor human cells and in a rat cell line transformed with the human oncogene. The promoter region is hypersensitive to DNAse I, micrococcal nuclease, endogenous nucleases and to S1 nuclease in supercoiled plasmids. This hypersensitive region is present in the different cell types analyzed and both normal and mutant alleles exhibit similar general sensitivity to DNAse I digestion in the same tumor cells. However, the 5' more distal DNAse I hypersensitive site, which is coincident with a region of the gene containing sequence homologies with known enhancers, exhibits variable sensitivity which appears to be higher in the tumor than in the normal and in the human than in the rat cells which we have analyzed. These data suggest the presence of specific factors interacting with the promoter sequences and delimits the transcription unit of the c-K-ras locus.
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PMID:Chromatin structure of the promoter region of the human c-K-ras gene. 376 6

As a step towards the understanding of possible relationship between chromatin organization and regulation of the oncogene expression, we have investigated the chromatin structure of one of the more frequently activated oncogenes, c-Ha-ras, in HeLa-S3 cells. This was accomplished by isolation of the chromatin fractions (soluble and insoluble) after micrococcal nuclease digestion of purified nuclei and probing for the distribution of ras sequences. The polynucleosomal fraction was further resolved by sucrose gradient sedimentation. Southern-blot hybridization of the DNA isolated from various fractions yielded following results: (1) c-Ha-ras sequences segregated predominantly in the lysate fraction. (2) Unlike the B-globin (transcriptionally inactive) sequences, ras-H associated chromatin lacked typical nucleosomal packaging. Furthermore, since post-translational modifications of nuclear proteins have been suggested to modulate the nucleosome structure during DNA transcription and replication, ras sequences, in polynucleosomes immunofractionated on anti-poly (ADP-Ribose) Sepharose were also examined. The data suggested that the major class of this oncogene sequence exists in chromatin more distal to the sites of this particular chromatin modification.
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PMID:The association of human c-Ha-ras sequences with chromatin and nuclear proteins. 388 46

A biosynthetic method has been developed that makes possible the site-specific incorporation of a large number of amino acids and analogues within proteins. In this approach, an amber suppressor tRNA chemically aminoacylated with the desired amino acid incorporates this amino acid site specifically into a protein in response to an amber codon introduced at the corresponding position in the protein's DNA sequence. Using this method, precise changes within a protein can be made to address detailed structure-function questions. A series of fluorinated tyrosine analogues and linear, branched, and cyclic hydrophobic amino acids have been used to determine the impact of hydrogen bonding and hydrophobic packing, respectively, on protein stability. Glutamate analogues and conformationally restricted amino acids have been used to probe the mechanisms of staphylococcal nuclease and ras. In addition, this technique has been used to construct photocaged proteins and proteins containing photoaffinity labels, spin labels, and isotopic labels at specific positions in the protein sequence suitable for biophysical studies.
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PMID:Site-directed mutagenesis with an expanded genetic code. 766 23

The activated c-Ha-rasVal12 oncogene is often involved in the genesis of human malignancies. We show here that in c-Ha-rasVal12 oncogene-transformed mouse NIH 3T3 fibroblasts the copy number and expression level of the mutant ras oncogene correlates with the degree of chromatin decondensation, as assessed by micrococcal nuclease (MNase) and DNase I digestion. MNase and DNase I analyses further revealed that the nucleosomal repeat lengths were different in the normal and ras oncogene-transformed cells, 162.3 bp and 178.1 bp, respectively. These chromatin changes were accompanied by alterations in the content of histone H1 zero. Furthermore, using DNase I as a probe, we discovered that serum stimulation of normal and transformed cells, synchronized by serum starvation, induces rapid reversible changes in the structure of bulk chromatin that may be linked to transcriptional activation. Our data thus indicate that cell transformation by ras is associated with specific changes in chromatin structure that make it more vulnerable, and prone to additional mutations characteristic of cancer development in vivo.
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PMID:Cell transformation by c-Ha-rasVal12 oncogene is accompanied by a decrease in histone H1 zero and an increase in nucleosomal repeat length. 772 50

Flavivirus infection causes extensive proliferation and reorganization of host cell membranes to form specialized structures called convoluted membranes/paracrystalline arrays and vesicle packets (VP), the latter of which is believed to harbor flaviviral replication complexes. Using detergents and trypsin and micrococcal nuclease, we provide for the first time biochemical evidence for a double membrane compartment that encloses the replicative form (RF) RNA of the three pathogenic flaviviruses West Nile, Japanese encephalitis, and dengue viruses. The bounding membrane enclosing the VP was readily solubilized with nonionic detergents, rendering the catalytic amounts of enzymatically active protein component(s) of the replicase machinery partially sensitive to trypsin but allowing limited access for nucleases only to the vRNA and single-stranded tails of the replicative intermediate RNA. The RF co-sedimented at high speed from nonionic detergent extracts of virus-induced heavy membrane fractions along with the released individual inner membrane vesicles whose size of 75-100 nm as well as association with viral NS3 was revealed by immunoelectron microscopy. Viral RF remained nuclease-resistant even after ionic detergents solubilized the more refractory inner VP membrane. All of the viral RNA species became nuclease-sensitive following membrane disruption only upon prior trypsin treatment, suggesting that proteins coat the viral genomic RNA as well as RF within these membranous sites of flaviviral replication. These results collectively demonstrated that the newly formed viral genomic RNA associated with the VP are oriented outwards, while the RF is located inside the nonionic detergent-resistant vesicles.
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PMID:Architecture of the flaviviral replication complex. Protease, nuclease, and detergents reveal encasement within double-layered membrane compartments. 1270 Feb 32