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

Hybrid nucleases consisting of an oligonucleotide fused to a unique site on the relatively nonspecific phosphodiesterase staphylococcal nuclease have been shown to sequence specifically cleave DNA. We have introduced mutations into the binding pocket of the nuclease which lower the kcat/Km of the enzyme. Hybrid nucleases generated from these mutants sequence selectively hydrolyze single-stranded DNA in a catalytic fashion, and under a much wider range of conditions than was previously possible. One such hybrid nuclease (Y113A, K116C) was able to site selectively cleave single-stranded M13mp7 DNA (7214 nt), primarily at one phosphodiester bond. Another hybrid nuclease (Y113A, L37A, K116C) catalyzed the hydrolysis of a 78-nt DNA substrate with a kcat of 1.2 min-1 and a Km of 120 nM. The effects of variations in the length and sequence of the oligonucleotide binding region were examined, as were changes in the length of the tether between the oligonucleotide and the enzyme. Cleavage specificity was also assayed as a function of substrate DNA primary and secondary structure and added poly(dA).
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PMID:Generation of a catalytic sequence-specific hybrid DNase. 260 84

We have designed and studied antisense oligodeoxynucleotides (oligonucleotides; oligos) which we call 'pseudo-cyclic oligonucleotides' (PCOs). PCOs contain two oligonucleotide segments attached through their 3'-3'- or 5'-5'-ends. One of the segments of the PCO is an antisense oligo complementary to a target mRNA, and the other is a short protective oligo that is 5-8 nucleotides long and complementary to the 3'- or 5'-end of the antisense oligo. As a result of complementarity between the antisense and protective oligo segments, PCOs form intramolecular pseudo-cyclic structures in the absence of the target RNA. The antisense oligo segment of PCOs used for the studies described here is complementary to an 18-nucleotide-long site on the mRNA of the protein kinase A regulatory subunit RIalpha (PKA-RIalpha). Thermal melting studies of PCOs in the absence and presence of the complementary RNA suggest that the pseudo-cyclic structures formed in the absence of the target RNA dissociate, bind to the target RNA, and form heteroduplexes. The results of RNase H cleavage assays suggest that PCOs bind to complementary RNA and activate RNase H in a manner similar to that of an 18-mer conventional antisense PS-oligo. In snake venom (a 3'-exonuclease) or spleen (a 5'-exonuclease) phosphodiesterase digestion studies, PCOs are more stable than conventional antisense oligos because of the presence of 3'-3'- or 5'-5'-linkages and the formation of intramolecular pseudo-cyclic structures. PCOs with a phosphorothioate antisense oligo segment inhibited cell growth of MDA-MB-468 and GEO cancer cell lines similar to that of the conventional antisense PS-oligo, suggesting efficient cellular uptake and target binding. The nuclease stability studies in mice suggest that PCOs have higher in vivo stability than antisense PS-oligos. The studies in mice showed similar pharmacokinetic and tissue distribution profiles for PCOs to those of antisense PS-oligos in general, but rapid elimination from selected tissues.
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PMID:Pseudo-cyclic oligonucleotides: in vitro and in vivo properties. 1065 77

The properties of new chimeric oligodeoxynucleotides made of short sequences (tetramers, pentamers, octamers, and decamers) bridged by hexamethylenediol and hexaethylene glycol linkers have been investigated. These chimeric oligonucleotides showed an improved resistance toward snake venom 3'-phosphodiesterase, with an increased stability when a terminal 3'-3'-internucleotide phosphodiester bond is present. It also has been demonstrated that the hybrid complexes formed by bridged oligonucleotides and a complementary 20-mer RNA are able to elicit the activity of ribonuclease H (RNase H) from Escherichia coli. The substrate properties of chimeric oligonucleotides depend on the length of the oligonucleotide fragments bridged by linkers. Introduction of a nonnucleotide spacer into the native oligonucleotide only slightly hampers the extent of the RNA hydrolysis in the hybrid complexes, whereas a modification of the site of reaction is observed as a possible consequence of the steric disturbance due to the aliphatic linkers. Hence, these new chimeric oligonucleotides, namely, short oligonucleotide fragments bridged by nonnucleotide linkers, demonstrate a favorable combination of exonuclease resistance and high substrate activity toward RNase H. As a consequence, these chimeric oligonucleotides could be proposed as new, promising analogs to be used in the antisense strategy.
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PMID:Nuclease resistance and RNase H sensitivity of oligonucleotides bridged by oligomethylenediol and oligoethylene glycol linkers. 1133 43

Fun30 is an ATP-dependent chromatin remodeler in budding yeast that is involved in cellular processes important for maintaining genomic stability such as gene silencing and DNA damage repair. Cells lacking Fun30 are moderately sensitive to the topoisomerase inhibitor camptothecin and exhibit a delay in cell cycle progression in the presence of camptothecin. Here, we show that Fun30 is required to cope with torsional stress in the absence of Top1. Moreover, we show through genetic studies that Fun30 acts in a parallel pathway to Mus81 endonuclease but is epistatic to Tdp1 phosphodiesterase and Rad1 endonuclease in the repair of camptothecin-induced DNA damage. More importantly, we show that DNA damage sensitivity of Fun30 deficient cells is enhanced in the absence of RNase H enzymes that remove RNA:DNA hybrids. We believe that chromatin remodeling by Fun30 may be important in dealing with torsional stress and camptothecin-induced DNA damage.
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PMID:Fun30 chromatin remodeler helps in dealing with torsional stress and camptothecin-induced DNA damage. 3314 48

The primary role of apurinic/apyrimidinic (AP) endonuclease APE1 in human cells is the cleavage of the sugar phosphate backbone 5' to an AP site in DNA to produce a single-strand break with a 5'-deoxyribose phosphate and 3'-hydroxyl end groups. APE1 can also recognize and incise some damaged or modified nucleotides and possesses some minor activities: 3'-5' exonuclease, 3'-phosphodiesterase, 3'-phosphatase, and RNase H. A molecular explanation for the discrimination of structurally different substrates by the single active site of the enzyme remains elusive. Here, we report a mechanism of target nucleotide recognition by APE1 as revealed by the results of an analysis of the APE1 process involving damaged DNA and native RNA substrates with non-canonical structures. The mechanism responsible for substrate specificity proved to be directly related to the ability of a target nucleotide to get into the active site of APE1 in response to an enzyme-induced DNA distortion.
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PMID:Activity of Human Apurinic/Apyrimidinic Endonuclease APE1 Toward Damaged DNA and Native RNA With Non-canonical Structures. 3319 55