Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.26.4 (RNase H)
 2,751 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Nuclease-resistant alpha-anomeric DNA:beta-RNA hybrids are inhibitors of Escherichia coli RNase H, and Drosophila embryo RNase H. RNase H activities were measured by polyacrylamide gel electrophoresis, employing a short substrate, (A)12:d[G-G-(T)12-G-G], or by acid-solubility techniques, using a long substrate, poly(A):poly(dT). Strand exchanges which could be responsible for the observed inhibition have been ruled out by S1 nuclease experiments and by using inhibitors which do not allow strand exchange. Our results suggest that RNase H, for which DNA:RNA duplexes are the natural substrates, binds to non-physiological alpha-DNA:RNA hybrids and is consequently inhibited. These hybrids also inhibit the RNA-dependent DNA polymerase activity of M-MLV reverse transcriptase, therefore appearing as potential inhibitors of at least two reverse transcriptase activities. However, the inhibitory effect of these hybrids with respect to M-MLV reverse transcriptase is also observed with the single-stranded alpha-DNA itself. Unexpectedly, polymerase activity is highly stimulated by alpha-oligos, analogous in their sequence to the beta primer used at a concentration unable to generate a detectable synthesis. These results suggest that the inhibition of reverse transcriptase activity with the alpha:beta may occur at different levels.
Gene 1988 Dec 10
PMID:Alpha-anomeric DNA: beta-RNA hybrids as new synthetic inhibitors of Escherichia coli RNase H, Drosophila embryo RNase H and M-MLV reverse transcriptase. 246 72

The reverse transcriptase (RT) activity of human immunodeficiency virus type 1 and other retroviruses is closely associated with a hybrid-degrading RNase H activity which is essential for retroviral replication. We have analyzed the effect of sulfated polysaccharides on human immunodeficiency virus type 1 recombinant RT and RNase H activities in vitro. Heparin, dextran sulfates, and xylan polysulfate were found to be much more potent inhibitors of RNase H than of RT and exhibit 50% infective doses of 0.04 to 0.1 micrograms/ml (corresponding to 0.1 to 25 nM) which is up to 5,000-fold more efficient than that for RT. Inhibitors of RNase H activity are attractive as antiviral drugs.
J Virol 1989 Dec
PMID:Inhibition of human immunodeficiency virus type 1 RNase H by sulfated polyanions. 247 74

A system for the functional reconstitution of yeast U2 snRNPs using synthetic U2 RNAs is described. We use oligonucleotide-directed RNase H cleavage to specifically deplete yeast extracts of their endogenous full-length U2 snRNA and consequently inactivate pre-mRNA splicing activity. The subsequent addition of synthetic yeast U2 RNAs, derived by in vitro transcription (T7U2 RNAs), to these oligonucleotide-treated extracts efficiently reconstitutes their ability to splice pre-mRNA. The use of deletion derivatives of the T7U2 RNA has demonstrated that the region downstream from the conserved Sm-binding site sequence in the yeast U2 RNA is not absolutely required for pre-mRNA splicing activity in vitro. Furthermore, we found that both human and rat U2 RNAs can function in yeast extracts. We also show that point mutations in the yeast U2 RNA can be analyzed using the in vitro reconstitution system. Allele-specific suppression of mutations in pre-mRNA branch site sequence is observed when the appropriate compensatory mutations in the branch site recognition region of the T7U2 RNA are introduced. Finally, we present a model for the interaction of the U2 and U6 snRNAs during pre-mRNA splicing.
Genes Dev 1989 Dec
PMID:In vitro reconstitution of functional yeast U2 snRNPs. 256 Jul 54

U6 small nuclear RNA (snRNA) is the most highly conserved spliceosomal RNA, and it has been postulated to have a fundamental role in pre-mRNA splicing. To elucidate this role, we developed an in vitro system for reconstituting the functional U6 small ribonucleoprotein (snRNP). Treating splicing extracts with an oligonucleotide complementary to the central domain of U6 snRNA leads to both RNase H cleavage of the endogenous U6 snRNA and loss of splicing activity. Yeast U6 RNA, synthesized in vitro using T7 RNA polymerase, is then added to the oligonucleotide-treated extract, and restoration of splicing activity is monitored by the subsequent addition of substrate pre-mRNA. Addition of full-length, unmodified T7U6 snRNA (113 nucleotides) to oligonucleotide-treated extracts restores splicing activity efficiently. Using U6 RNA transcripts truncated at their 3' ends, we show that large deletions (39 nucleotides) produce molecules that are unable to restore splicing activity in vitro and cannot interact with the endogenous U4 snRNA or form a mature spliceosome. Finally, we show that substitution of the invariant G81 with C within the T7U6 RNA abolishes its ability of restoring splicing activity. Although the U4/U6 snRNP forms correctly, mature spliceosomes do not assemble.
Genes Dev 1989 Dec
PMID:In vitro assembly of yeast U6 snRNP: a functional assay. 256 Jul 55

Previous studies have revealed multiple size classes of rat insulin-like growth factor-I (IGF-I) of estimated size 7.5-7.0, 1.9-1.5, and 1.2-0.9 kilobases (kb). Available sequence information accounts for only 2.1 kb of the 7.5-7.0 kb IGF-I mRNAs. We used oligomer directed ribonuclease H (RNase H) mapping to define the extent to which the unknown sequence in the large molecular weight mRNAs lies 5' or 3' to known sequence. Rat liver polyadenylated RNAs were incubated with oligomer probes complementary to internal rat IGF-I precursor (E domain) coding sequences. RNase H was used to hydrolyze IGF-I mRNAs at the point of annealment with the oligomers. Resultant 5' and 3'-IGF-I mRNA fragments were analyzed on Northern blots. A probe specific for type 1 (class C) 5'-sequences (the most predominant of multiple 5'-sequence types found on rat IGF-I mRNAs) identifies intact IGF-I mRNAs of 7.5-7.0, 1.9-1.5 and 1.2-0.9 kb but, after oligomer directed RNase cleavage of these mRNAs, identified only a single IGF-I mRNA 5'-fragment. Major differences in the length of sequence 5' to the IGF-I coding sequence therefore, do not account for the multiple size classes of type 1 (class C) IGF-I mRNAs. The size of the 5'-fragment suggests that the extent of sequence 5' to the IGF-I coding sequence is 0.4-0.7 kb in type 1 (class C) IGF-I mRNAs. Identification of multiple 3'-fragments of IGF-I mRNAs demonstrated heterogeneity in the 3'-ends of rat IGF-I mRNAs.(ABSTRACT TRUNCATED AT 250 WORDS)
Mol Endocrinol 1989 Dec
PMID:The size heterogeneity of rat insulin-like growth factor-I mRNAs is due primarily to differences in the length of 3'-untranslated sequence. 256 Aug 8

We have located a positive, cis-acting DNA sequence element within the 5' flanking DNA of the c-myc gene (-125 base pairs). This DNA sequence element has a large purine-pyrimidine strand asymmetry and can assume the H-DNA conformation. A factor with the properties of a ribonucleoprotein (RNP) interacts with this DNA region. The interaction of the c-myc DNA sequence element and the RNP involves an RNase H-sensitive mechanism and, therefore, may involve an RNA.DNA hybrid. In addition, a protein factor(s) binds to this DNA sequence element. DNA footprinting and mutant oligonucleotide binding/competition assays implicate a punctate, poly(G.C) recognition/binding sequence for the RNP factor, whereas the major protein factor requires two ACCCT sequence motifs for maximal binding. These results suggest that RNP and protein factors act as positive transcriptional regulators of the c-myc gene, perhaps by altering DNA topology.
Proc Natl Acad Sci U S A 1989 Dec
PMID:Ribonucleoprotein and protein factors bind to an H-DNA-forming c-myc DNA element: possible regulators of the c-myc gene. 269 70

The replication of simian virus 40 origin-containing DNA has been reconstituted in vitro with SV40 large T antigen and purified proteins isolated from HeLa cells. Covalently closed circular DNA (RF I') daughter molecules are formed in the presence of T antigen, a single-stranded DNA binding protein and DNA polymerase alpha-primase complex, together with ribonuclease H, DNA ligase, topoisomerase II, and a double-stranded specific exonuclease that has been purified to homogeneity. The 44-kDa exonuclease-digested oligo(rA) annealed to poly(dT) in the 5'----3' direction. DNA ligase and the 5'----3' exonuclease were essential for RF I' formation. Covalently closed circular duplex DNA and full length linear single-stranded DNA were detected by alkaline gel electrophoresis as products of the complete system. DNA replication in the absence of either DNA ligase or the 5'----3' exonuclease yielded DNA products that were half length (approximately 1500 nucleotides) and smaller Okazaki-like fragments (approximately 200 nucleotides). Hybridization experiments showed that the longer chains were synthesized from the leading strand template, while the small products were synthesized from the lagging strand template. These results suggest that the RNA primers attached to 5' ends of replicated DNA are completely removed by the 5'----3' exonuclease, with the assistance of RNase H.
J Biol Chem 1988 Dec 25
PMID:Complete enzymatic synthesis of DNA containing the SV40 origin of replication. 284 39

A series of antisera directed against amino acid sequences from different segments of the duck hepatitis B virus (DHBV) P-gene were shown to immunoprecipitate DHBV DNA molecules that were covalently linked to the DHBV DNA terminal protein. Restriction analysis and sizing after protease treatment demonstrated that the P-gene proteins were bound to the 5'-end of the DHBV DNA minus-strand which was mapped to a G-residue in the centre of the repeat sequence DR1. Resistance to alkali treatment indicated a phosphodiester linkage to tyrosine between protein and DNA. Limited protease treatment prior to immunoprecipitation cleaved C-terminal P-proteins from the viral DNA, indicating that the terminal protein forms a separate domain encoded in the N-terminal part of the P-gene. Functional analysis of a deletion mutant confirmed the notion that a non-essential spacer separates the terminal protein from the polymerase domain residing in the C-terminal half of the P-gene. Thus, the major proteins required for hepadnaviral reverse transcription, namely the primer, DNA polymerase, and possibly also RNase H, appear to be synthesized as a polyprotein precursor which is at least initially linked as such to its first DNA product.
EMBO J 1988 Dec 20
PMID:The amino-terminal domain of the hepadnaviral P-gene encodes the terminal protein (genome-linked protein) believed to prime reverse transcription. 285 56

T helper cells have recently been divided into two subsets. The Th1 subset secretes and responds to IL-2 in an autocrine manner. The Th2 subset upon mitogen or antigen stimulation releases IL-4. Here we describe a novel technology that allowed us to confirm this distinction. We have used synthetic oligonucleotides complementary to the 5' end of mouse IL-2 and IL-4 to specifically block the biosynthesis of IL-2 or IL-4 in two murine helper T cell clones from the Th1 or Th2 subset. We show that the antisense IL-2 oligonucleotide inhibited the proliferation of the Th1 clone and had no effect on the Th2 clone. In parallel experiments, the antisense IL-4 oligonucleotide blocked the proliferation of the Th2 clone and not the proliferation of the Th1 clone. The inhibition was significantly reversed in both cases by the addition of the relevant lymphokine (IL-2 in the case of the Th1 clone, IL-4 in the case of the Th2 clone). Northern analysis, using cDNA probes specific for the two lymphokines, showed a decrease in the steady-state level of the relevant lymphokine mRNA, suggesting the specific degradation of the mRNA by an RNase H-like enzymatic activity. This strategy, which allows the specific blockade of the biosynthesis of a lymphokine, could be useful for future studies on the role of each T helper subset in physiological immune responses.
J Exp Med 1988 Dec 01
PMID:Specific inhibition of lymphokine biosynthesis and autocrine growth using antisense oligonucleotides in Th1 and Th2 helper T cell clones. 297 66

A number of enzymes thought to be involved in DNA replication have been identified in the brain. These include single-stranded DNA-binding proteins, topoisomerases I and II, DNA polymerase alpha, a protein that binds Ap4A and might be classified as a DNA polymerase alpha accessory protein, RNase H, DNA polymerase beta, DNA ligase, an endo- and an exonuclease of unknown function, DNA methyl transferase and poly(ADPR) synthase. In contrast, little is known about the enzymology of DNA repair in brain. The few enzymes identified comprise uracil-DNA glycosylase, DNA polymerase beta, DNA polymerase alpha (which in neurons is present only at immature stages), DNA ligase, poly(ADPR) synthase, and O6-alkylguanine-DNA alkyltransferase. In addition, an exonuclease acting on depurinated single-stranded DNA (tentatively listed here as 3'----5' exonuclease), an endonuclease of unknown function as well as ill-defined acid and alkaline deoxyribonucleases also occur in brain.
Brain Res 1985 Dec
PMID:Enzymology of DNA replication and repair in the brain. 300 64


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