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)

Pyran copolymer (maleic anhydride-divinyl ether) has consistently reproducible molecular weight-related biologic effects associated with toxic, immunologic antitumor, and antiviral effects. Fortunately, the antitumor action occurs with the least toxic lower molecular weight fraction. Immunoadjuvant effects with this fraction would be critical to its development. Studies of polymers should include evaluation of effects on splenomegaly, splenic esterase changes, lipolysis, reverse transcriptase, nucleases, calcium flux, cyclic nucleotides, and complement and clotting elements.
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PMID:Future direction of synthetic polyanions (pyran copolymer). 36 28

A chick non-muscle alpha-actinin cDNA probe encoding the EF-hand region of molecule was used to screen a lambda gt10 chick brain cDNA library from 14-day embryos. A partial 2.1-kb alpha-actinin cDNA was isolated (8W cDNA) which encoded a protein identical to chick skeletal-muscle alpha-actinin, except in the C-terminal part of the first EF hand. In the variant, the 22 residues found in the skeletal-muscle isoform were replaced by a stretch of 26 unique residues. Analysis of the structure of the skeletal-muscle alpha-actinin gene showed that the region of divergence was encoded by two exons which are alternatively spliced. Quantitative reverse transcriptase/polymerase chain reaction (RT/PCR) was used to investigate the levels of the alpha-actinin transcripts in various tissues. The skeletal-muscle alpha-actinin variant was expressed at low levels in brain, liver and spleen, but could not be detected in skeletal muscle. Surprisingly, skeletal-muscle alpha-actinin mRNA was also expressed in brain, liver and spleen. The RT/PCR products were authenticated by using diagnostic restriction enzyme sites and by sequencing. The splice variant derived from the skeletal-muscle alpha-actinin gene was also detected in a variety of cDNA libraries from both adult and embryonic tissues by PCR. Although a transcript encoding this alpha-actinin splice variant is expressed in non-muscle tissues, neither of the two EF-hands would be predicted to be functional, making it unlikely to be a typical non-muscle isoform which are calcium-sensitive with respect to binding actin. The two vertebrate non-muscle alpha-actinins sequenced to date also have a spacer of five amino acids between the two EF hands, whereas in the variant, the spacer is just four residues in length. Further analysis will be required before this alpha-actinin isoform, which we refer to as SKv, can be classified as muscle or non-muscle alpha-actinin. We propose a new nomenclature to describe the various alpha-actinin genes and their transcripts.
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PMID:A chick skeletal-muscle alpha-actinin gene gives rise to two alternatively spliced isoforms which differ in the EF-hand Ca(2+)-binding domain. 148 65

Human immunodeficiency virus type 1 (HIV-1) integrase (IN) is the viral protein required for integration of the HIV-1 genome into host cell DNA. A series of clones expressing portions of IN as lambda cII fusion proteins has been constructed in an Escherichia coli expression system; a Southwestern procedure was used to examine binding of the expressed proteins to DNA oligonucleotides. Proteins expressed by clone pHIP106, encoding the entire IN protein but no other pol sequence, and pKNA101, which expresses an IN fusion protein containing 23 amino acids of HIV-1 reverse transcriptase at its amino terminus, exhibited similar levels of oligonucleotide binding. Little DNA sequence specificity was associated with binding activity and there was a preference for Mn2+ over Mg2+ and Ca2+. Interestingly, the protein expressed by an N-terminal clone containing nucleotides coding for IN amino acids 1-141 (including a conserved His-Cys box) was unable to bind oligonucleotide, whereas the protein expressed by a C-terminal clone containing nucleotides coding for amino acids 142-288 exhibited binding equivalent to that of full-length IN. The C-terminal protein was unreactive with a MAb to the lambda cII leader peptide and with an antipeptide serum directed against amino acids 141-158. These results are consistent with the previously reported internal initiation of IN protein synthesis in E. coli at met 154, and indicate that the C-terminal clone does not express IN amino acids 142-153. These amino acids represent part of a conserved region termed D(35)E, containing amino acids 116-152, which has been implicated in IN DNA binding.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Localization of DNA binding activity of HIV-1 integrase to the C-terminal half of the protein. 154 Apr 16

Retroviral RNA is copied into DNA by reverse transcriptase when the viral genome enters into its life cycle. In the case of human immunodeficiency virus (HIV), massive amounts of unintegrated viral DNA reportedly appear in the early phase of primary infection. However, the relationship between the accumulation of this DNA and the cytopathic effect (CPE) remains obscure. In an attempt to delineate this association, we examined the appearance of the unintegrated viral DNA by means of two experimental systems: (1) primary infection of highly susceptible MOLT-4#8 cells and (2) induction of CPE by cell-fusion of persistently infected MOLT-4#8 cells. A correlation was observed between the accumulation of unintegrated viral DNA and the appearance of CPE, both when MOLT-4#8 cells were infected with cell-free virus and when persistently infected MOLT-4#8 cells were co-cultured with uninfected cells. Persistently infected cells did not fuse spontaneously in culture, because they lack the CD4-molecule on their surfaces. However, when treated with polyethylene glycol (PEG), the cells fused, exhibited ballooning degeneration, and released fewer viruses. After PEG treatment, unintegrated viral DNA also appeared. Since such DNA is generally not detected in persistently infected cells, it is possible that some cellular mechanism exists to suppress the synthesis of viral DNA and that the fusion induced by PEG treatment cancels the suppression. Treatment of persistently infected cells with Ca2+ ionophore and Ca2+ antagonist also resulted in the accumulation of unintegrated viral DNA and inhibited virus release. These findings suggest that the induction of unintegrated HIV DNA may be an effective strategy for reducing the release of the virus.
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PMID:Unintegrated DNA in cells infected in vitro with human immunodeficiency virus (HIV): a new approach to suppression of virus release. 169 87

The DNA polymerase and RNase H activities of HIV reverse transcriptase are both essential for HIV replication. Although the two activities are both catalyzed by a single polypeptide, they are physically separate; i.e., the DNA polymerase resides in the N-terminal domain whereas the RNase H is localized in the C-terminal domain. The present study was undertaken to characterize the enzymatic properties of these two activities and to determine whether the two catalytic sites are also functionally distinct. We have observed that EGTA specifically stimulates, whereas CaCl2 selectively inhibits, the RNA-dependent DNA polymerase activity but that neither compound has any effect on the RNase H activity of a recombinant HIV reverse transcriptase. The stimulation of the DNA polymerase activity by EGTA is dependent on the Mg2+ concentration; the greatest stimulation is observed at low Mg2+ concentrations. Similarly, the inhibition of DNA polymerase activity by Ca2+ is influenced by Mg2+ concentration. Ca2+ inhibition can be reversed by increasing Mg2+ concentrations, suggesting the possibility that CaCl2 inhibits the reverse transcriptase activity by competing for a metal-binding site on the enzyme. The pyrophosphate analogue phosphonoformate selectively inhibits the polymerase activity but not the RNase H activity of HIV reverse transcriptase. In contrast, the RNase H activity can be selectively inhibited by deoxyadenosine 5'-monophosphate, whereas the DNA polymerase activity is not inhibited. These results suggest that the DNA polymerase and RNase activities are not only physically separate but that they are also functionally distinct.
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PMID:Functional characterization of RNA-dependent DNA polymerase and RNase H activities of a recombinant HIV reverse transcriptase. 170 16

The pyrophosphate analogue, foscarnet, selectively inhibits the DNA polymerase of human herpes viruses, including cytomegalovirus, and the reverse transcriptase of HIV. Viral replication is therefore prevented, but resumes when the drug is cleared from infected cells. In vitro, the combination of foscarnet and zidovudine (azidothymidine) has an additive effect against cytomegalovirus and acts synergistically against HIV. An improvement in cytomegalovirus retinitis is obtained in over 85% of affected AIDS patients during foscarnet induction therapy, but relapse usually occurs within a month of ceasing treatment. There is a similar duration of remission during maintenance therapy given for 5 days each week, but this can be extended 4- to 5-fold with daily administration of higher doses. In allograft recipients, progression of retinitis can be halted by foscarnet until immune function recovers and eradicates the virus. The incidence of acute renal failure, which is common during foscarnet therapy, may be reduced by dosage adjustment and adequate prehydration. Anaemia, phlebitis, nausea and vomiting, and disturbances in serum calcium and phosphate levels, perhaps resulting from uptake of foscarnet into bone or chelation with ionised calcium, have also been associated with administration of the drug. Cytomegalovirus retinitis is difficult to treat, with few therapeutic options available. Although treatment with foscarnet produces some severe adverse effects, with care these can be minimised, and the drug produces clinical improvement in a large proportion of patients; this is a highly encouraging finding at this stage in its development. Preliminary comparative data indicate that foscarnet and ganciclovir are similarly effective, but foscarnet may have some theoretical advantages in AIDS patients since it can be used in combination with zidovudine without potentiating myelosuppression.
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PMID:Foscarnet. A review of its antiviral activity, pharmacokinetic properties and therapeutic use in immunocompromised patients with cytomegalovirus retinitis. 170 82

Culture supernatants from the rabbit macrophage cell line 6083 infected with a retrovirus, human immunodeficiency virus type 1 (HIV-1), were negative for reverse transcriptase (RT) expression although the line was shown to be productively infected by all other criteria tested. Supernatants from uninfected cultures of 6083, the human monocyte line U937, and from freshly isolated peripheral human monocytes, were found to contain a monocyte-derived inhibitory factor (MDIF) which interferes with a standard assay for RT. MDIF is a heat-labile activity of approximately of 40 kD. Both substrates and products of the reverse transcriptase assay are degraded by MDIF which is not affected by reduction and alkylation of disulfide bonds. MDIF is inhibited by the addition of a particular thioated oligonucleotide (S-dG30) to the reaction mixture but this addition also inhibits RT. The optimum method to minimize MDIF interference in the RT assay is by addition of ethylene glycol bis-(beta-aminoethyl ether)N,N,N',N'-tetraacetic acid (EGTA); MDIF requires divalent cations for activity and has a strong preference for calcium which is preferentially chelated by EGTA. The potential presence of this inhibitory activity should be considered when using RT levels as a measure of retroviral infection.
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PMID:A monocyte-derived factor interferes with detection of reverse transcriptase in HIV-1 infection. 170 43

In characterizing a group of independent human immunodeficiency virus (HIV-1) isolates, we noted that certain isolates had anomolously low levels of virion-associated reverse transcriptase activity. In an attempt to understand the basis of this phenomenon, we examined in detail one such isolate, HIV-1G. We found correctly processed forms of the viral reverse transcriptase in virions as well as processed forms of other viral proteins, suggesting that viral proteins are both expressed and properly processed. We have detected a nuclease activity associated with the outer face of the HIV-1G envelope. This nuclease degrades the DNA product generated during the reverse transcription assay. The nuclease activity is more sensitive to mild protein denaturation than is the viral reverse transcriptase, and it is stimulated by the presence of Ca2+. The amount of virion-associated nuclease activity relative to reverse transcriptase activity varies between virus isolates and can vary also for one isolate during virus spread through a culture. The origin of the nuclease activity is unknown but is presumed to be cellular. The variability in amount of nuclease activity may reflect variability in the interaction of the virus with different cellular components during maturation.
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PMID:Characterization of an HIV-1 isolate displaying an apparent absence of virion-associated reverse transcriptase activity. 171 16

To elucidate the action of vitamin C on pathogenic human retroviruses, we investigated and compared the effects of noncytoxic concentrations of ascorbic acid (AA), its calcium salt (Ca-ascorbate), and two thiol-based reducing agents [glutathione (GSH) and N-acetyl-L-cysteine (NAC)] against human immunodeficiency virus (HIV)-1 replication in chronically infected T lymphocytes. Ca-ascorbate reduced extracellular HIV reverse transcriptase (RT) activity by about the same magnitude as the equivalent dose of AA. Long-term experiments showed that continuous presence of ascorbate was necessary for HIV suppression. NAC (10 mmol/L) caused less than twofold inhibition of HIV RT and conferred a synergistic effect (approximately eightfold inhibition) when tested simultaneously with AA (0.426 mmol/L). In contrast, nonesterified GSH (less than or equal to 1.838 mmol/L) had no effect on RT concentrations and did not potentiate the anti-HIV effect of AA. These results further support the potent antiviral activity of ascorbate and suggest its therapeutic value in controlling HIV infection in combination with thiols.
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PMID:Comparative study of the anti-HIV activities of ascorbate and thiol-containing reducing agents in chronically HIV-infected cells. 172 May 98

A new Amphotericin B derivative, MS-8209, which retains high antifungal activity with greatly reduced toxicity and improved solubility, has been developed. We investigated the antiviral properties of MS-8209 in Jurkat and CEM T-cell lines and in peripheral blood mononuclear cells infected in vitro with HIV-1BRU. Our results demonstrate, by determination of reverse transcriptase activity and p24 antigen level titration in cell culture supernatants, that MS-8209 inhibits HIV-1 replication in all cell types at concentrations without cytotoxicity. MS-8209 also prevents membrane expression of the HIV-1 large envelope glycoprotein gp120 and the decrease in CD4 level at the surface of infected cells. HIV-1-infected Jurkat cells exhibit a severe signalling defect at CD3 stimulation. Treatment with MS-8209 restores normal responsiveness at CD3 as assessed by measurement of inositol triphosphate accumulation and calcium flux. Finally, our results indicate that MS-8209 inhibits HIV-1BRU replication without preventing virus binding and penetration into target cells.
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PMID:MS-8209, a new Amphotericin B derivative that inhibits HIV-1 replication in vitro and restores T-cell activation via the CD3/TcR in HIV-infected CD4+ cells. 172 40


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