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)

The interactions between beef tRNATrp with avian myeloblastosis reverse transcriptase have been studied by statistical chemical modifications of phosphate (ethylnitrosourea) and cytidine (dimethyl sulfate) residues, as well as by digestion of complexed tRNA by Cobra venom nuclease and Neurospora crassa endonuclease. Results with nucleases and chemicals show that reverse transcriptase interacts preferentially with the D arm, the anticodon stem and the T psi stem. All these regions are located in the outside of the L-shaped structure of tRNA. This domain of interaction is different to that reported previously in the complex of beef tRNA with the cognate aminoacyl-tRNA synthetase (M. Garret et al.; Eur. J. Biochem. In press). Avian reverse transcriptase destabilizes the region of tRNA where most of the tertiary interactions maintaining the structure of tRNA are located.
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PMID:Interactions between avian myeloblastosis reverse transcriptase and tRNATrp. Mapping of complexed tRNA with chemicals and nucleases. 620 Aug 30

A convenient and rapid technique for preparing radiolabeled single-stranded DNA hybridization probes has been developed. Single-stranded recombinant M13 phage DNA containing the mRNA strand of a cloned cDNA is bound to diazobenzyloxymethyl-cellulose in a manner that permits the synthesis of a complementary DNA using reverse transcriptase and primed with either oligo(dT) or the M13 single-stranded primer. A procedural advantage is that after synthesis the unincorporated radiolabeled nucleotides are washed away easily, and the radiolabeled single-stranded DNA probe is eluted with formamide, ready for use. To limit the DNA copy to the insert, a preliminary synthesis reaction is performed with unlabeled nucleotides, primer, and enzyme, followed by digestion of the reaction mix with a restriction endonuclease that recognizes a unique site in the recombinant immediately upstream of the cDNA insert. After elution of the unlabeled synthesized complementary DNA, a second synthesis reaction yields highly radiolabeled single-stranded DNA that extends only the length of the mRNA insert. A major advantage is that the restriction enzyme-cleaved, cellulose-bound template can be stored and reused repeatedly.
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PMID:Synthesis of single-stranded hybridization probes from reusable DNA templates bound to solid support. 620 47

The nucleotide sequence of the coding portion of human alpha globin mRNA has been determined by sequence analysis using human alpha globin cDNA cloned in bacterial plasmids. The sequence was obtained by a combination of direct sequence analysis of the cloned cDNA and analysis of cDNA obtained by primer extension, using short restriction endonuclease fragments of cloned alpha cDNA that were hybridized to human globin mRNA and elongated on the mRNA template by viral reverse transcriptase. The human alpha globin mRNA has an unexpectedly high G + C base composition (64.7%), similar to that observed for rabbit globin alpha mRNA, and displays a striking bias in the use of synonym codons for various amino acids. The bias in codon usage of human alpha globin mRNA is similar, with some exceptions, to that previously observed for rabbit alpha globin mRNA as well as for human and rabbit beta globin mRNAs. A detailed restriction endonuclease map of the human alpha globin cDNA is presented.
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PMID:Nucleotide sequence of the coding portion of human alpha globin messenger RNA. 624 94

We have previously described a nonconditional mutant of avian sarcoma virus (SE21Q1b) which fails to package viral RNA (Gallis et al., Virology 94:146-161, 1979; Linial et al., Cell 15:1371-1381, 1978). Quail cells transformed by SE21Q1b contain normal amounts of intracellular viral mRNA's for src, env, and gag-pol and release particles with the density of normal virus containing a typical complement of virion proteins, including reverse transcriptase. These virions are noninfectious for both chicken and quail cells and contain primarily cellular rather than viral RNA. Analysis by gel electrophoresis of the cellular DNA of quail cells transformed by SE21Q1b after restriction endonuclease digestion indicated the presence of a single provirus. The provirus was located at one site in the genome of the host cell and was flanked by the characteristic terminally repeated sequences derived from the 3' and 5' ends of viral RNA. The only defect detected in the SE21Q1b provirus was a deletion of ca. 150 base pairs of DNA somewhere between 300 and 600 bases from the left (gag-pol) end of the provirus. Analyses of the proviral DNA of cells transformed by wild-type recombinants between SE21Q1b and leukosis viruses reveal that the recombinants no longer contain this deletion. The deletion, therefore, defines a region on the viral RNA which is required for correct packaging of the virion RNA.
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PMID:Avian oncovirus mutant (SE21Q1b) deficient in genomic RNA: characterization of a deletion in the provirus. 625 70

Preparations of purified Rauscher murine leukemia virus were found to contain an endodeoxyribonuclease after disruption of the virus with nonionic detergents. The enzyme makes single-strand breaks in linear or covalently closed circular phage double-stranded DNA molecules. The enzyme was partially purified by ion-exchange chromatography on DEAE- and carboxymethyl-Sepharose columns followed by electrophoresis in DNA-containing polyacrylamide gels. The enzyme was separated from reverse transcriptase (p80pol), and the final endonuclease preparation contained no detectable reverse transcriptase activity. The DEAE-Sepharose column-purified endonuclease activity contained a polypeptide of about 40,000 Mr that we term p40. Peptide mapping experiments demonstrated that p40 shares methionine-labeled tryptic peptides with Pr200gag-pol and Pr135pol. Six major methionine-labeled tryptic peptides derived from p40 were found in Pr200gag-pol, but only five of these were detected in Pr135pol. The four core proteins (p30, p15, pp12, and p10) and p80pol plus p40 account for most, but not all, of the peptide sequences of Pr200gag-pol. The endonuclease-associated p40 is similar in size and precursor origin to the avian retrovirus-coded endonuclease (p32). In view of these similarities to the avian p32 endonuclease and its association with partially purified Rauscher murine leukemia virus-associated endonuclease preparations, we propose that p40 is the Rauscher murine leukemia virus-coded endonuclease.
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PMID:Endodeoxyribonuclease activity associated with Rauscher murine leukemia virus. 626 Sep 82

Chromosome-mediated transfer of murine leukaemia (MuLV) and murine sarcoma (MuSV) virus genetic information to uninfected recipient cells was investigated. Metaphase chromosomes from AKR MuLV-infected SC-1 mouse cells were incubated with NIH/3T3 cells. After several passages (1 to 3 weeks), infectious virions exhibiting reverse transcriptase activity and the characteristic host range of ecotropic, N-tropic AKR virus appeared in the supernatant fluids of the treated cells. Restriction endonuclease analysis of genomic DNA from transfected cells indicated that AKR proviral DNA was associated with the high molecular weight DNA of the host. These results demonstrate that the AKR MuLV genome can be stably transferred to uninfected recipient cells via isolated metaphase chromosomes. Although AKR virions are not able to infect heterologous cells, chromosome-mediated transfection resulted in the establishment of productive AKR MuLV infection in mink cells. Thus, the use of chromosomes to transfer virus genes can circumvent the natural host restriction barrier. In other experiments, it was shown that normal NIH/3T3 cells were transformed after exposure to metaphase chromosomes isolated from an MuSV-infected, non-producer line. Foci were detected 14 to 21 days after chromosome treatment and were shown to contain true viral transformants since transforming virus was produced after superinfection with MuLV.
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PMID:Transfer of murine leukaemia and murine sarcoma virus genetic information by transfection with isolated metaphase chromosomes. 629 50

A new cell line designated SQ-A was established from the spleen of a leukemic DBA/2J mouse inoculated with the anemic strain of Friend erythroleukemia virus (FLV-A). The cells are similar in morphology, growth pattern, and tumorigenicity to our prototype erythroleukemia line 5-86 but are more sensitive to the cytotoxic effects of inducers of differentiation. The virus produced by SQ-A cells induces erythroleukemia associated with anemia in adult mice but has little activity when assayed on XC cells. It was characterized to determine what factors influence its leukemogenic potential. As compared to the attenuated virus from cultures of 5-86 and G-2 cells, the subunits of the RNA from the virions of SQ-A cells are the same size, and the amount of reverse transcriptase activity and RNase H present in the purified virions of the three lines are similar. However, differences are observed in levels of endonuclease and protein kinase. Both enzymes are increased in SQ-A virions. The activity of protein kinase in SQ-A virions is about 5 times higher than that in the attenuated virions. The number of polypeptides and their phosphorylation patterns also distinguish the virions of SQ-A. Whereas 5-86 virions contain seven proteins, three of which are phosphorylated in vitro, SQ-A virions contain eight proteins, all of which are phosphorylated. The extra protein in SQ-A virions has a molecular weight of 25,000 and is not glycosylated.
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PMID:Characterization of leukemogenic virus produced by a new line of Friend erythroleukemia virus-transformed cells. 632 17

Calmodulin mRNA has been partially purified from a total nucleic acid extract of the electroplax of Electrophorus electricus by oligo(dT)-cellulose chromatography and sucrose gradient centrifugation. A 9- to 10S fraction was determined to contain 39% calmodulin mRNA by translation in a reticulocyte lysate followed by immunoprecipitation with antibodies to calmodulin. Double-stranded cDNA was synthesized from the RNA fraction by using reverse transcriptase from avian myeloblastosis virus. The double-stranded cDNA was joined to pBR322 linearized by restriction endonuclease Pst I and used to transform Escherichia coli RRI. DNAs from 60 tetracycline-resistant cloned hybridized to [32P]cDNA synthesized from the partially purified calmodulin mRNA fraction. By direct DNA sequence analysis, one of these clones, pCM109, was shown to contain calmodulin-specific sequences corresponding to amino acid residues 93--148 of calmodulin or approximately 38% of the peptide-coding region of the calmodulin structural gene sequence. pCM109 was hybridized to DNA isolated from three vertebrate and one plant species by the procedure of Southern. Positive hybridization bands were noted regardless of the DNA source. These data suggest thaat calmodulin gene sequences are evolutionarily conserved, as has been shown to be the case for the primary amino acid sequence.
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PMID:A cloned calmodulin structural gene probe is complementary to DNA sequences from diverse species. 694 Dec 92

Cultures of murine Friend erythroleukemia (FL) cells, which are chronically infected with leukemia virus, were inoculated with vaccinia virus. The yield of vaccinia virus was determined by assaying plaque-forming units in mouse L2 cells, and the yield of leukemia virus was determined by measuring reverse transcriptase (RNA-dependent DNA nucleotidyltransferase) activity released into the culture fluid. Although no facilitation of one virus by the other was detected, persistently infected cultures were established. Electron microscopic examination revealed the presence of vaccinia and leukemia viruses in the same cell. The permanent lines of cells persistently infected with vaccinia were designated FLvac. Their morphology, growth rate, cloning efficiency, and ability to respond to the induction of erythrodifferentiation by treatment with dimethyl sulfoxide were not appreciably altered as compared to the parental FL cells. However, the persistently infected cells showed a marked decrease in tumorigenicity when assayed in DBA/2 mice. The infectious virus produced by FLvac cells and by L2 cells were indistinguishable as judged by restriction endonuclease patterns of virion DNA, structural proteins, and the activities of two virion-associated DNases. The yield of infectious vaccinia virus from FLvac cells generally declined after about 60 serial passages. Although some cell lines no longer yield infectious virus, they are resistant to challenge with vaccinia at concentrations that are cytolytic for L2 cells. The mechanism responsible for the establishment of the persistent infection remains unclear because defective particles, interferon production, and temperature-sensitive mutants have not been detected.
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PMID:Persistent infection of Friend erythroleukemia cells with vaccinia virus. 695 93

Influenza viral RNA transcription in vitro is primed by capped RNA fragments cleaved from capped RNAs by a viral endonuclease. The present study was undertaken to determine whether the specificities of the viral endonuclease and transcriptase observed in in vitro studies are also observed in the infected cell. The NS (nonstructural) gene of influenza WSN virus was cloned in pBR322 by using a double-stranded DNA containing a cDNA copy of both virion RNA (vRNA) and in vivo viral mRNA. We determined the 5' terminal sequence of the particular NS viral mRNA molecule which was cloned and also the 5' terminal sequences of the entire population of in vivo NS viral mRNAs synthesized in two different cell lines. For the latter determination we used a restriction fragment from the cloned DNA for the reverse transcriptase-catalyzed extension of total in vivo viral mRNA. The results indicate that in vivo and in vitro viral RNA transcription are similar in two important respects: (i) transcription initiates not with an A residue directed by the 3' terminal U of the vRNA, but with a G residue directed by the 3' penultimate C of the vRNA; and (ii) capped RNA fragments containing a 3' terminal A residue are preferentially used as primers, therapy generating an AG sequence in the viral mRNA complementary to the 3' terminal UC of the vRNA. Actually, for in vivo transcription, a subset of A-terminated capped fragments, namely those containing a 3' penultimate C residue, are the preferred primers. The latter specificity had not been observed in previous in vitro studies.
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PMID:Selected host cell capped RNA fragments prime influenza viral RNA transcription in vivo. 730 81


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