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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)

Many crystal forms of human immunodeficiency virus reverse transcriptase have been obtained by vapour diffusion, microbatch and microdialysis methods. Despite their apparent morphological perfection, no X-ray diffraction has been discernible in most cases with these crystals.
J Mol Biol 1991 Jan 05
PMID:Many crystal forms of human immunodeficiency virus reverse transcriptase. 170 35

The gene coding for starch phosphorylase (EC 2.4.1.1) was isolated from a potato genomic library constructed in lambda EMBL3. It is an unusually long plant gene (16.4 kb) which encodes a preprotein of 966 amino acids. The phosphorylase coding sequence is interrupted by 14 introns whose positions do not match those of the introns in the human glycogen phosphorylase gene. A 78 amino acid central peptide unique to plant plastidial phosphorylases is hypothesized to have arisen through the mis-splicing of an intron-exon junction site in an ancestral gene. The fifth intron of the phosphorylase is very large (approximately 7 kb) and contains a copia-like transposable element inserted in the opposite orientation to that of the phosphorylase gene. This element has been named Tst1; it is bordered on the 5' and 3' sides by long terminal repeats of 285 and 283 bp respectively, which define an internal domain of 4492 bp. Tst1 contains 4 open reading frames (ORFs) that encode protein domains for a reverse transcriptase, an integrase, an RNA-binding site and a protease. Transcription of the phosphorylase gene appears to proceed unimpaired through the copia element.
Mol Gen Genet 1990 Oct
PMID:Occurrence of a copia-like transposable element in one of the introns of the potato starch phosphorylase gene. 170 27

The LaBelle-1b strain of Neurospora intermedia contains a 4.1-kb closed-circular mitochondrial plasmid DNA, which encodes a single long open reading frame of 1,151 amino acids reported to have sequence similarity to reverse transcriptases. Here, we show that the LaBelle strain contains a novel DNA polymerase activity that is highly specific for the endogenous LaBelle plasmid DNA in nucleoprotein particles and can be distinguished from the mitochondrial DNA polymerase by several characteristics. Photolabeling experiments indicate that the LaBelle-specific DNA polymerase activity is associated with a polypeptide of 120 kDa, which is in good agreement with the size predicted for the protein encoded by the LaBelle plasmid open reading frame (132 kDa). This 120-kDa polypeptide is found only in the LaBelle strain that contains the mitochondrial plasmid, and it cosegregates with mitochondria in sexual crosses, suggesting that it is encoded by the plasmid. The LaBelle-specific DNA polymerase efficiently uses the artificial DNA substrates, poly(dA)-oligo(dT) and poly(dC)-oligo(dG), but despite its reported sequence similarity to reverse transcriptases, it has very low activity with analogous RNA substrates, poly(rA)-oligo(dT), poly(rC)-oligo(dG), or poly(rCm)-oligo(dG). Considered together with the previous sequence comparisons, our results suggest that the LaBelle plasmid encodes a novel DNA polymerase, which was derived from a protein that was at one time a reverse transcriptase but lost its ability to use RNA templates. This DNA polymerase now presumably functions in replication of the plasmid. Our results constitute the first biochemical evidence for a DNA polymerase activity associated with a mitochondrial plasmid. Further, they may provide insight into the evolution of DNA polymerases from reverse transcriptases, as presumably occurred in the course of evolution following the transition from the so-called RNA world to the present DNA world.
Mol Cell Biol 1991 Mar
PMID:The LaBelle mitochondrial plasmid of Neurospora intermedia encodes a novel DNA polymerase that may be derived from a reverse transcriptase. 170 12

A genetic element, called a retron, is present in certain Escherichia coli strains. It consists of genes for the production of a covalently linked DNA-RNA compound and a reverse transcriptase. The presence of a retron can be detected by testing for a satellite DNA band by polyacrylamide gel electrophoresis. This DNA band consists of the DNA portion of the DNA-RNA compound and is called msDNA (multicopy single-stranded DNA). In a survey of intestinal E. coli isolates we detected msDNAs in classical enteropathogenic (EPEC) strains and in strains with aggregative adherence to tissue-culture cells (AA), but not in enteroinvasive (EIEC) and enterotoxigenic (ETEC) strains. Among 76 EPEC strains belonging to 14 different serotypes, msDNA was found to be present in 7 serotypes. In total, five different types of msDNA were found, although within each serotype, the msDNAs were the same. These results suggest that different retrons are clonally inherited.
Mol Microbiol 1990 Oct
PMID:Distribution of msDNAs among serotypes of enteropathogenic Escherichia coli strains. 170 55

The partial sequences of 16S rRNA from Mycoplasma bovis and M. agalactiae were determined by dideoxynucleotide sequencing using reverse transcriptase. Two oligonucleotides complementary to different evolutionary variable regions of 16S rRNA from these two species were synthesized. The oligonucleotides were end-labelled with 32P and used as probes in filter hybridization experiments with different bovine, caprine and ovine mycoplasmas as samples. One of the probes, complementary to a sequence of the V8-region of both M. bovis and M. agalactiae, did not cross-hybridize to any bovine, caprine or ovine mycoplasmas except M. bovigenitalium and M. californicum. This probe is thus not useful for analysis of bovine samples, but can be used for detection of M. agalactiae in samples from goats and sheep, since M. bovigenitalium and M. californicum have never been isolated from these hosts and M. bovis only occasionally. The other probe, complementary to a sequence of the V6-region of M. bovis, gave some cross-hybridization with M. agalactiae but not with bovine mycoplasmas. M. agalactiae has never been isolated from cattle and this probe is therefore useful for rapid screening of bovine samples for M. bovis.
Mol Cell Probes 1991 Feb
PMID:Detection of Mycoplasma bovis and Mycoplasma agalactiae by oligonucleotide probes complementary to 16S rRNA. 170 7

An msDNA operon, consisting of genes for msDNA and a reverse transcriptase, is present in Escherichia coliB and absent from E. coliK12. We have found that the msDNA operon is located on a DNA fragment, longer than 15kb, that is absent from E. coliK12. Using conjugation, P1 transduction, and nucleic acid hybridization between E. coliB and E. coliK12 strains, we have located the position of the msDNA operon on the E. coliB chromosome at a site that corresponds to minute 19 on the genetic map and to position 900 on the physical map of the E. coliK12 chromosome.
Mol Microbiol 1990 Dec
PMID:Mapping of the msDNA operon in the chromosome of Escherichia coli B. 170 40

In the search for novel derivatives of 1-[2-(hydroxyethoxy)methyl]-6-(phenylthio)thymine (HEPT), we have found that several 5-ethyl-6-(3,5-dimethylphenylthio)uracil and 5-ethyl-6-(3,5-dimethylbenzyl)uracil analogues are exquisitely potent and selective inhibitors of human immunodeficiency virus type 1 (HIV-1) replication in a variety of cell culture systems. Of this series, 5-ethyl-1-ethoxymethyl-6-(3,5-dimethylbenzyl)uracil (E-EBU-dM) emerged as the most active congener. Its 50% inhibitory concentration for HIV-1 (HTLV-IIIB) in MT-4 cells and peripheral blood lymphocytes is 2.2 and 0.45 nM, respectively. These concentrations are more than 10(5)-fold lower than the 50% cytotoxic concentrations of E-EBU-dM for the host cells. All compounds proved equally inhibitory to a number of clinical HIV-1 isolates, including a 3'-azido-2',3'-dideoxythymidine-resistant variant. However, as previously noted for HEPT, they do not inhibit human immunodeficiency virus type 2 replication. Reverse transcriptase assays have revealed that these HEPT derivatives act specifically on HIV-1 reverse transcriptase, according to a mechanism that is different from that of the dideoxynucleosides.
Mol Pharmacol 1991 Jun
PMID:Highly potent and selective inhibition of human immunodeficiency virus type 1 by a novel series of 6-substituted acyclouridine derivatives. 171 Nov 48

The role of cytokines in vivo has been difficult to assess. This difficulty is due, in part, to the limited number of producer cells and the strict regulation of cytokine production. In order to address this situation, we have developed assays which allow us to quantitate both protein production and steady state mRNA levels from specific in vivo sites. In this report, we present data utilizing these assays on cells obtained from draining LN following specific sensitization with antigen in vivo. In order to determine the relative quantities of cytokine mRNA, we modified the reverse transcriptase-polymerase chain reaction which had been previously described. The modified assay is (1) linear over a large concn range of input template (2) demonstrates a high degree of reproducibility (SE approximately 13%) and (3) is very sensitive. Utilizing this assay, we have measured a constitutive mRNA (DHFR), quantitated both the presence of lymphokine mRNA (IL-2) and the induction of cytokine mRNA (TNF alpha). In this report we have examined the kinetics of TNF alpha mRNA expression and have demonstrated that following epicutaneous sensitization with picryl chloride, there is rapid induction (within 24 hr) of TNF alpha mRNA in the draining LN and that the levels of mRNA remain detectable through d7. In addition, we determined the time course of production of TNF protein by the draining LN cells and found that it was similar to that of the mRNA levels. A potential pathologic role for immune response generated TNF alpha is also discussed. We believe these experiments demonstrate that cytokine production following antigen-specific sensitization in vivo can be analyzed at both the cellular and molecular level. The data suggests that this approach can be used to study cytokine regulation in vivo.
Mol Immunol
PMID:Quantitation of cytokine mRNA levels utilizing the reverse transcriptase-polymerase chain reaction following primary antigen-specific sensitization in vivo--I. Verification of linearity, reproducibility and specificity. 171 71

The recombinant reverse transcriptase of HIV-1 virus has been isolated from Escherichia coli cells transformed by the plasmid pRT40 DNA. The 103 Kd protein produced by these cells is shown to be processed to proteins with lower molecular masses by the reverse transcriptases own protease activity as well as Escherichia coli proteases. The resulting 103-66 Kd proteins possess the polymerase activity while 51 Kd and smaller proteins are lacking the activity. The 66 and 51 Kd reverse transcriptase fragments demonstrate the positive immunological reaction with the human blood serum from the people possessing antibodies to HIV-1 virus. The recombinant reverse transcriptase of HIV-1 produced by Escherichia coli cells is shown to be useful in AIDS diagnosis in humans.
Mol Gen Mikrobiol Virusol 1991 Mar
PMID:[Polymerase and immunologic activity of reverse transcriptase of the HIV-1 virus, isolated from Escherichia coli]. 171 97

The human immunodeficiency virus 1 (HIV-1) reverse transcriptase (RT) is a protein of 66 kDa, p66, which contains two domains, an amino-terminal DNA polymerase and an RNase H at the carboxy terminus of the molecule. In order to characterize the mode of action of the RNase H, two previously described mutant enzymes were used, with substitutions in the highly conserved histidine 539, which was mutated to the neutral amino acid asparagine and to the negatively charged aspartate. The purified wild-type (wt) and mutant (mt) enzyme activities are analyzed here using RNA-DNA hybrids consisting of in vitro transcribed RNA that harbors the polypurine tract (PPT) from HIV-1 and DNA oligonucleotides complementary to the PPT or to other regions of the RNA. Analysis of the radioactively labeled RNA of these model hybrids after RNase H treatment indicates that both, wt and mt enzymes, are capable of cleaving the RNA in an endonucleolytic manner. The mt enzymes exhibit a severely reduced exonuclease activity. They are more sensitive towards salt and competition with excess of unlabeled hybrid, suggesting a reduced substrate binding affinity. DNA elongation by the RT is coupled with RNA hydrolysis by the 3'-5' exonuclease of the wt RNase H. The RNase Hmt of the mt enzymes, however, does not exhibit such processive 3'-5' exonuclease activity during DNA synthesis but gives rise to sporadic endonucleolytic cuts, whereas the RT is not affected. The endonuclease activities of the RNase H mt enzymes exhibit cleavage preferences in the absence or presence of DNA synthesis different from those of the wt enzyme. They cannot recognize specific sequences required to generate a PPT-primer and therefore cannot initiate plus-strand DNA synthesis in vitro at the 3' end of the PPT, which is essential for viral replication.
J Mol Biol 1991 Aug 05
PMID:Mutations of a conserved residue within HIV-1 ribonuclease H affect its exo- and endonuclease activities. 171 5


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