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:3.1.27.5 (RNase)
17,967 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The physiologic substrates of cytotoxic T lymphocyte granule-associated serine esterases (referred to hereafter as proteases or "granzymes"), and the role of these enzymes in cell-mediated activity remain unclear. We have developed an assay for possible ligands of the trypsin-like dimeric serine protease granzyme A based on Western immunoblotting techniques. This protein-binding assay demonstrates the selective binding of granzyme A to several proteins present in the target cell P815. The binding specificity is preserved when enzyme binding is performed in the presence of excess competing proteins, including such cationic species as lysozyme and RNase. Enzyme binding is inhibited, however, by heat or detergent inactivation of granzyme A. Subcellular fractionation of target cells shows that the nuclear fraction contains most granzyme A binding reactivity, which is recovered in the nuclear salt wash fraction. A protein with Mr = 100,000 and two closely migrating proteins with Mr = 35,000 and 38,000 are the predominant reactive moieties, and the N-terminal sequence of the 100-kDa protein confirmed that this protein was murine nucleolin. Incubation of granzyme A with nucleolin generates a discrete proteolytic cleavage product of Mr = 88,000. Since nucleolin is known to shuttle between nucleus and cytoplasm, the interaction of granzyme A and nucleolin may be important in the process of apoptosis which accompanies cytotoxic T lymphocyte-mediated lysis of target cells.
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PMID:Granzyme A binding to target cell proteins. Granzyme A binds to and cleaves nucleolin in vitro. 186 Aug 69

When fixed preparations of newt germinal vesicle (GV) contents are treated with RNase and are then probed with radiolabeled single-stranded DNA in 0.1-2.0 X SSC, the extrachromosomal nucleoli bind the probe non-specifically. DNA/protein blot analysis of proteins from newt GVs shows that gv95, an acidic protein (pI = 5.0) of Mr = 95,000, is the most prominent non-specific DNA-binding protein. Immunocytochemical analysis with affinity purified antibody directed against gv95 shows that it is located in the multiple nucleoli. We used an antibody directed against rat nucleolin to show that newt gv95 and two similar Xenopus GV proteins are the amphibian versions of nucleolin, a nucleolar ribonucleoprotein originally identified in mammalian cells. We show that mAb 3A10, directed against newt histones H1 and H5, labels gv95 on protein immunoblots and the multiple nucleoli in cytological preparations. These results suggest that histone H1 and nucleolin share a cross-reacting epitope.
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PMID:Nucleolin from the multiple nucleoli of amphibian oocyte nuclei. 219 42

In ATT, a human autoimmune serum, we found anti-nucleolar antibodies that recognized nucleolar antigens confined to a single nucleolar compartment, the dense fibrillar component (DFC). We localized these antigens by immunoelectron microscopy in DFC of HeLa cell nucleoli both on Lowicryl sections and cryoultrathin sections without embedding. The antigens were solubilized by incubation with 2M NaCl but not by RNase or DNase treatment. The ATT serum crossreacted with rat liver nucleoli and PtK1 cell nucleoli in which immunofluorescence labelling displayed a clumpy pattern. During mitosis, the antigens dispersed in the cytoplasm until late telophase, when they gathered in the prenucleolar bodies. In human peripheral lymphocytes, or HeLa cells treated with actinomycin D, the antigens were still present but the fluorescence intensity decreased. By immunoblotting using human nuclear extracts, the ATT serum bound to a 116,000 Mr protein at dilutions up to 1:2000. The reactivity of this band diminished with actinomycin D-treated nuclear extracts. Two minor bands were also observed at 97 and 70K (K = 10(3) Mr). Immunopurification by competition or elution demonstrated that the 116K antigens were at the origin of the nucleolar labelling. This DFC marker appeared to be different from the NOR-silver-stained proteins, which in our preparations exhibited apparent molecular weights of 105, 80 and 38-40K. In addition, these 116K antigens did not exhibit the characteristics described for DNA topoisomerase I, fibrillarin or nucleolin. We propose the 116K antigen as a new marker of the DFC of the nucleoli.
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PMID:A 116,000 Mr nucleolar antigen specific for the dense fibrillar component of the nucleoli. 220 Jul 92

Acholeplasma laidlawii A has been grown in media containing synthetic, long chain C20- and C23-fatty acids possessing a diacetylene group in their acyl chains. Growth on the C23 diacetylenic acid was poor but was good on the C20 acid. Biosynthetic incorporation of the fatty acids occurs; as much as 90% of the membrane lipid fatty acyl chains consisting of the C20-diacetylenic fatty acid, the remainder being shorter chain, saturated fatty acids. The thermal phase transition of this biomembrane has been studied and a differential scanning calorimetry heating curve shows the presence of an endotherm corresponding to a membrane lipid phase transition occurring at about 26 degrees C. The lipid class composition of membranes containing the C20-diacetylene lipids was examined and found to be similar to membranes from cells grown on oleic acid-containing medium. (The ratio of monoglucosyl- to diglucosyldiacylglycerols was the same but the ratio of glycolipid to phosphatidylglycerol was higher in the cells grown with diacetylene fatty acids). Upon irradiation with ultraviolet light the cells and isolated biomembranes become coloured, either red or yellow depending upon their thermal history. The colour change indicates that extensive cross-linking of the lipids of the biomembranes of A. laidlawii has occurred and that a conjugated polymeric structure has been formed. Analysis of the extracted lipids from the biomembranes by GLC indicates that extensive cross-linking of the lipid chains within the biomembrane of a natural cell system has been achieved. The monoglucosyldiacylglycerols cross-link more readily that do the phosphatidylglycerol lipids. The effect of such lipid cross-linking or polymerisation on the activity at 35 degrees C of an intrinsic membrane-bound enzyme, NADH oxidase, and ribonuclease, an extrinsic membrane-bound enzyme, was studied. The NADH oxidase activity decreased rapidly upon cross-linking of the lipid environment whereas ribonuclease activity was unaffected. The potential for future studies of polymerised model and natural biomembranes is discussed.
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PMID:The biosynthetic incorporation of diacetylenic fatty acids into the biomembranes of Acholeplasma laidlawii A cells and polymerisation of the biomembranes by irradiation with ultraviolet light. 683 76

Most DNA topoisomerase II (topo II) in cell-free extracts of 0-2-h old Drosophila embryos appears to be nonnuclear and remains in the supernatant after low-speed centrifugation (10,000 g). Virtually all of this apparently soluble topo II is particulate with a sedimentation coefficient of 67 S. Similar topo II-containing particles were detected in Drosophila Kc tissue culture cells, 16-19-h old embryos and extracts of progesterone-matured oocytes from Xenopus. Drosophila topo II-containing particles were insensitive to EDTA, Triton X-100 and DNase I, but could be disrupted by incubation with 0.3 M NaCl or RNase A. After either disruptive treatment, topo II sedimented at 9 S. topo II-containing particles were also sensitive to micrococcal nuclease. Results of chemical cross-linking corroborated those obtained by centrifugation. Immunoblot analyses demonstrated that topo II-containing particles lacked significant amounts of lamin, nuclear pore complex protein gp210, proliferating cell nuclear antigen, RNA polymerase II subunits, histones, coilin, and nucleolin. Northern blot analyses demonstrated that topo II-containing particles lacked U RNA. Thus, current data support the notion that nonnuclear Drosophila topo II-containing particles are composed largely of topo II and an unknown RNA molecule(s).
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PMID:An RNase-sensitive particle containing Drosophila melanogaster DNA topoisomerase II. 808 68

Nuclei assembled in Xenopus egg extract contain numerous spherical aggregations or nuclear bodies. Previously we have shown that they closely resemble prenucleolar bodies (PNBs), both at the compositional and ultrastructural level. Subsequently, coilin was also identified and for this reason they were called coiled bodies. Here we present morphological and immunocytochemical evidence that the in vitro nuclear bodies resemble authentic PNBs and are different from coiled bodies. In particular we show that coilin, previously considered as the defining protein constituent of coiled bodies, is also present in PNBs of cultured cells. In contrast, the PNB-associated nucleolar proteins nucleolin and B23/NO38 are not detectable in coiled bodies and may thus serve as suitable markers for PNBs. Our results suggest that PNBs are primary assembly structures which contribute to the formation of both nucleoli and coiled bodies and thus offer an explanation for the frequently observed structural association of coiled bodies with nucleoli. To gain some insight into the assembly process of PNBs in vitro, specific nucleolar proteins were removed from Xenopus egg extract. Quite surprisingly, the immuno-depleted extracts still promoted the assembly of nuclear bodies which lacked either fibrillarin, nucleolin, xNopp180 or B23/NO38. Only after fibrillarin-depletion fewer PNBs were seen as compared to controls. Digestion of the extract with RNase followed by northern blot analysis revealed that U3 small nucleolar RNA is not required for the formation and structural maintenance of PNBs in vitro.
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PMID:Prenucleolar bodies contain coilin and are assembled in Xenopus egg extract depleted of specific nucleolar proteins and U3 RNA. 901 Jul 83

Monoclonal antibodies binding to different domains of nucleolin have been used to localize nucleolin in tissue culture cells of Xenopus laevis. The monoclonal antibody b6-6E7 binds to an epitope in the N-terminal domain, which contains arrays of phosphorylation consensus sites. This monoclonal antibody binds to nucleolin of oocytes and of eggs with high affinity. In contrast, the monoclonal antibody Nu-1H6 binds poorly to the modified forms of nucleolin arising during meiosis and mitosis. In interphase cells, monoclonal antibody b6-6E7 preferentially stains the periphery of the nucleoli, where most of the rRNA accumulates. Staining by monoclonal antibody Nu-1H6 complements this pattern by staining mainly the center of the nucleoli. The epitope of monoclonal antibody Nu-1H6 is within the central domain of nucleolin, which contains the first two RNA binding domains. RNase treatment of cells results in loss of nucleolin from nucleoli. In mitotic cells, both monoclonal antibodies decorate the surface of condensing chromosomes in prophase. The periphery of the condensed chromosomes in metaphase and anaphase is preferentially stained by monoclonal antibody b6-6E7.
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PMID:Subcellular distribution of distinct nucleolin subfractions recognized by two monoclonal antibodies. 952 40

Previous studies showed that components implicated in pre-rRNA processing, including U3 small nucleolar (sno)RNA, fibrillarin, nucleolin, and proteins B23 and p52, accumulate in perichromosomal regions and in numerous mitotic cytoplasmic particles, termed nucleolus-derived foci (NDF) between early anaphase and late telophase. The latter structures were analyzed for the presence of pre-rRNA by fluorescence in situ hybridization using probes for segments of pre-rRNA with known half-lives. The NDF did not contain the short-lived 5'-external transcribed spacer (ETS) leader segment upstream from the primary processing site in 47S pre-rRNA. However, the NDF contained sequences from the 5'-ETS core, 18S, internal transcribed spacer 1 (ITS1), and 28S segments and also had detectable, but significantly reduced, levels of the 3'-ETS sequence. Northern analyses showed that in mitotic cells, the latter sequences were present predominantly in 45S-46S pre-rRNAs, indicating that high-molecular weight processing intermediates are preserved during mitosis. Two additional essential processing components were also found in the NDF: U8 snoRNA and hPop1 (a protein component of RNase MRP and RNase P). Thus, the NDF appear to be large complexes containing partially processed pre-rRNA associated with processing components in which processing has been significantly suppressed. The NDF may facilitate coordinated assembly of postmitotic nucleoli.
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PMID:Partially processed pre-rRNA is preserved in association with processing components in nucleolus-derived foci during mitosis. 972 3

rRNA precursors are bound throughout their length by specific proteins, as the pre-rRNAs emerge from the transcription machinery. The association of pre-rRNA with proteins as ribonucleoprotein (RNP) complexes persists during maturation of 18S, 5.8S, and 28S rRNA, and through assembly of ribosomal subunits in the nucleolus. Preribosomal RNP complexes contain, in addition to ribosomal proteins, an unknown number of nonribosomal nucleolar proteins, as well as small nucleolar RNA-ribonucleoproteins (sno-RNPs). This report describes the use of a specific, rapid, and mild immunopurification approach to isolate and analyze human RNP complexes that contain nonribosomal nucleolar proteins, as well as ribosomal proteins and rRNA. Complexes immunopurified with antibodies to nucleolin-a major nucleolar RNA-binding protein-contain several distinct specific polypeptides that include, in addition to nucleolin, the previously identified nucleolar proteins B23 and fibrillarin, proteins with electrophoretic mobilities characteristic of ribosomal proteins including ribosomal protein S6, and a number of additional unidentified proteins. The physical association of these proteins with one another is mediated largely by RNA, in that the complexes dissociate upon digestion with RNase. Complexes isolated from M-phase cells are similar in protein composition to those isolated from interphase cell nuclear extracts. Therefore, the predominant proteins that associate with nucleolin in interphase remain in RNP complexes during mitosis, despite the cessation of rRNA synthesis and processing in M-phase. In addition, precursor rRNA, as well as processed 18S and 28S rRNA and candidate rRNA processing intermediates, is found associated with the immunopurified complexes. The characteristics of the rRNP complexes described here, therefore, indicate that they represent bona fide precursors of mature cytoplasmic ribosomal subunits.
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PMID:Association of nonribosomal nucleolar proteins in ribonucleoprotein complexes during interphase and mitosis. 988 Mar 28

We investigated the interaction of a highly potent acridine-based tat-antagonist with the TAR RNA of HIV-1. The wild type TAR RNA and three mutants with U-->C23, G x C-->C x G26-39 or G x C-->A x U26-39 substitutions were used as substrates to study the molecular basis of drug-TAR RNA complex formation. Melting temperature and RNase protection experiments reveal that the G x C26-39 pair is a critical element for specific major groove recognition of TAR at the pyrimidine bulge. The results provide a rational basis for future design of optimized tat/TAR inhibitors.
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PMID:Molecular basis of HIV-1 TAR RNA specific recognition by an acridine tat-antagonist. 1042 76


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