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:2.7.7.49 (reverse transcriptase)
31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

These studies were designed to determine if RIDP was present in a particulate fraction of brains from patients with ALS and PD. Evidence that we have detected RIDP is as follows: (a) DNA polymerase activity persists in the presence of concentrations of actinomycin D and distamycin that inhibit most DNA-directed DNA synthesis (25); (b) the majority of endogenous DNA polymerase activity is sensitive to prior treatment with RNase; (c) the early reaction product is a 4-5 S DNA heteropolymer joined by hydrogen bonds to an RNA molecule; and (d) the purified [3H]DNA product anneals to RNA extracted from the enzyme-containing pellet more extensively than to normal brain RNA or poly(rA). The enzyme activity is in a cytoplasmic particle that can be sedimented at high speed and has the buoyant density of RNA tumor viruses (1.16-1.18 gm/ml). This particulate fraction is not disrupted by physical manipulation and maintains its characteristic density with repeated centrifugations. Treatment with the nonionic surfactant Sterox changes the buoyant density of the enzyme-containing particle to 1.24 gm/ml, the density of the onconavirus virion core. Synthesis of RNA-DNA hybrids by an endogenous reverse transcriptase reaction was found only in normal and diseased Chamorro brains. Examination of a limited number of normal and diseased brains from individuals who lived in the United States produced negative results (39). Definitive characterization of this polymerase activity and identification as a true viral polymerase will depend on purification of biochemically active quantities of this polymerase to determine its template specificities, its cation preference, the fidelity of its transcription product, as well as its antigenic relationship to animal virus and human leukemic RIDP. Of critical importance in these studies will be differentiation of this activity from normal brain DNA polymerase gamma and terminal deoxynucleotidyltransferase.
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PMID:RNA tumor viruses as causative agents of chronic neurological disease. 6 87

Conditions have been determined under which reverse transcriptase catalyzes the synthesis of the high yields of full length complementary deoxyribonucleic acid (cDNA). These conditions depend not only on the cencentration of deoxynucleoside triphosphates (1) but also on the concentration of reverse transcriptase. An analysis of the kinetics of cDNA synthesis and the size of cDNA synthesized as a function of time under different conditions indicates that the mechanism of action of reverse transcriptase is partially distributive. This accounts for the necessity of a high enzyme concentration to obtain high yields of full length cDNA. Additional experiments indicate that the yield of cDNA is limited by the fact that the template mRNA is rapidly inactivated. This is most likely due to the fact that the product cDNA is hydrogen bonded to the template mRNA during synthesis.
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PMID:The syntheiss of high yields of full-length reverse transcripts of globin mRNA. 7 63

The avian retrovirus RNA-directed DNA polymerase contains an activity that is capable of removing hydrogen bonds from duplex nucleic acid molecules. This "unwinding-like" activity appears to be specific in its action, affecting RNA.DNA and DNA.DNA duplex molecules but not RNA.RNA duplexes. Studies with defined RNA.DNA hybrid molecules (e.g., Rous sarcoma virus RNA and complementary DNAs representing specific regions of the Rous sarcoma virus genome) and DNA.DNA duplexes indicate that, although this activity can remove a portion of the hydrogen bonds from these double-stranded structures, complete separation of complementary strands is not accomplished. The unwinding-like activity exhibits sensitivities to temperature and monovalent and divalent cation concentrations. It can also remove a specific large oligonucleotide from the 5' end of the viral genome subsequent to RNase H hydrolysis of viral RNA complexed to DNA present at that terminus. This reverse transcriptase-associated unwinding-like activity is discussed with respect to recently proposed models of retrovirus proviral DNA synthesis.
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PMID:Unwinding-like activity associated with avian retrovirus RNA-directed DNA polymerase. 7 11

Extracts from over 100 normal human placentas have been examined for RNA-directed DNA polymerase (DNA nucleotidyltransferase, EC 2.7.7.7) activity. More than 80% of these placentas contained this enzyme activity, which banded at a density of 1.15-1.17 g/ml in sucrose. After heat treatment, this enzyme activity was shifted in density to 1.22-1.24 g/ml. The enzymatic activity was greater with (rC)n.(dG)12-18 than with (dC)n.(dG)12-18 and was not stimulated by (dG)12-18 alone. The product of the endogenous reaction, which was sensitive to RNase, had the characteristics of a small DNA associated with a large RNA by hydrogen bonding. Electron microscopic inspection of the material with a density of 1.15-1.17 g/ml revealed numerous retrovirus-like particles with central electron-dense cores and double-membraned envelopes. The enzyme may be associated with the retrovirus-lik particles noted in the trophoblast layer of some human placentas.
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PMID:Normal human placentas contain RNA-directed DNA polymerase activity like that in viruses. 8 52

In vitro transcription of the avian tumor virus RNA by RNA-directed DNA polymerase is initiated on the unique cellular 4S RNA. Previous studies have shown that on the average there is one such RNA primer hydrogen bonded to each viral 35S RNA. The present study confirms that finding and demonstrates that, at least for the majority of 35S RNA molecules, the primer is bound at a site close to the 5'-terminus.
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PMID:Site on the RNA of an avian sarcoma virus at which primer is bound. 16 90

The complete nucleotide sequence of the 5' noncoding region of the adenovirus 2 hexon messenger RNA has been established by sequence analysis of reverse transcripts. Such transcripts were generated by extension of specific single-stranded DNA primers with reverse transcriptase after hybridization to purified hexon mRNA. The total length of the 5' noncoding region was determined to be 240 nucleotides, of which the spliced tripartite leader sequence contributes 202 nucleotides including the terminal m7G. The sizes of the different segments of the tripartite leader were estimated by comparing the established mRNA sequence with the genomic sequences for the first and third leader segments, and were found to be 42 nucleotides for the first segment, 71 nucleotides for the second and 89 nucleotides for the third. The estimates are ambiguous, however, due to the presence of tandemly repeated sequences at both ends of the intervening sequence between the third leader segment and the body of the hexon mRNA. The sequence of the leader allows the formation of hydrogen-bonded interactions with the 3' end of 18S ribosomal RNA near the capped 5' end and also close to the initiator AUG.
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PMID:Sequence analysis of adenovirus DNA: complete nucleotide sequence of the spliced 5' noncoding region of adenovirus 2 hexon messenger RNA. 45 52

The recent finding that 3'-amino-3'-deoxythymidine 5'-triphosphate is a noncompetitive inhibitor of the HIV-1 reverse transcriptase (Kedar, P.S.; et al. Biochemistry 1990, 29, 3603-3611), prompted an investigation of the conformation of 3'-amino-3'-deoxythymidine. An X-ray diffraction study has revealed that the glycosidic torsion angle of the nucleoside is in the less common syn region and this solid-state geometry is stabilized by a three-dimensional network of self-associated hydrogen-bonded molecules. On the other hand, the aqueous solution conformation, as determined by 1H NMR, places the glycosidic torsion angle in the more usual anti region with the sugar in an equilibrium between C3'-endo and C2'-endo puckering. The energy barrier between the solid-state and solution conformation is relatively low as was demonstrated by the MM2 calculations.
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PMID:Solid-state and solution conformation of 3'-amino-3'-deoxythymidine, precursor to a noncompetitive inhibitor of HIV-1 reverse transcriptase. 171 58

An important aspect of the infection by the human immunodeficiency virus (HIV-1) type 1 is its clinical latency, suggesting that the virus itself or the provirus may remain latent for extended periods of time after primary infection. Certain heterologous viral proteins or chemical and physical agents are able to reactivate latent virus. Since a common denominator shared by these agents is the ability to cause stress response in cells, we have examined the effects of oxidative stress mediated by hydrogen peroxide (H2O2) on HIV-1 latently infected promonocytic cell line termed U1. After exposure to H2O2 in concentrations ranging from 0.1 to 2 mM, the viability of the U1 cells decreased during 24 h before recovery. At 24 h post stress, the U1 cells began to express virus as assessed by elevated reverse transcriptase activities in culture supernatants. Immunofluorescence carried out on stressed U1 cells using anti-HIV-1 polyclonal antibodies showed that H2O2 leads to HIV-1 gene expression activation, but not to a release of viral particles from damaged cells. Additionally, using a HeLa cell line containing integrated the bacterial chloramphenicol acetyl transferase (CAT) gene under the control of the HIV-1 long terminal repeat (LTR), we have shown that oxidative stress mediated by H2O2 allows transactivation of the viral LTR revealed by intracellular CAT activity. A stimulation factor of around 4 of CAT activity can be reached when these cells are treated with 0.5 mM H2O2.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Activation of human immunodeficiency virus type 1 by oxidative stress. 207 16

3-Methylthymine was synthesized into DNA copolymers and deoxynucleoside triphosphate to study its effect on DNA synthesis by the Klenow fragment of Escherichia coli polymerase I and avian myeloblastosis virus reverse transcriptase. Both polymerases were greatly inhibited by template 3-methylthymine. In response to 3-methylthymine, misincorporation of dTTP increased slightly, but occurred only at low levels consistent with spontaneous misincorporation in vitro. Surprisingly, template 3-methylthymine resulted in a striking decrease in background misincorporation, relative to normal incorporation by the Klenow fragment, of dGTP and, to a lesser extent, of dATP and dCTP. The incorporation of 3-methyl-dTTP into DNA was studied using DNA sequencing technology. The Klenow fragment failed to incorporate 3-methyl-dTTP even at 1 mM. Reverse transcriptase incorporated 3-methyl-dTTP opposite adenine, cytosine, and thymine, but at only about 1/40,000th the efficiency of complementary deoxynucleoside triphosphate incorporation. Furthermore, synthesis generally stalled at sites of 3-methyl-thymine incorporation. From these results, we conclude that damage at the central hydrogen-bonding position of thymine abolishes its base-pairing capabilities during DNA synthesis.
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PMID:DNA damage at thymine N-3 abolishes base-pairing capacity during DNA synthesis. 244 69

The crystal structure of 3'-azido-3'-deoxy-thymidine (AZT), an antiviral agent that inhibits HIV reverse transcriptase, has been determined from three-dimensional x-ray diffractometer data. The crystal structure contains two independent molecules of AZT forming a hydrogen bonded dimer but exhibiting different conformations. These conformations are different from those theoretically calculated by molecular mechanics methods. The azido groups associate with each other and interrupt the base stacking, forming a sandwich of two stacked bases. The close conformational similarity of AZT to thymidine explains why AZT is a good substrate for thymidine kinase. The selective inhibition of reverse transcriptase by AZT is not due to any conformational restrictions imposed by the azido group but likely due to their stereoelectronic properties.
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PMID:Conformation and sandwiching of bases by azido groups in the crystal structure of 3'-azido-3'-deoxy-thymidine (AZT), an antiviral agent that inhibits HIV reverse transcriptase. 245 15


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