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

---

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

In this study, we have located the sites of transcription initiation and termination on a cloned fragment of ribosomal DNA from X. laevis, and have sequenced the surrounding nucleotides. As reported previously (Reeder, Sollner-Webb and Wahn, 1977), about 25% of the 40S rRNA precursor molecules isolated from oocytes have polyphosphate 5' termini and are therefore presumed to represent primary transcripts. These ends hybridize specifically to the 221 bp DNA fragment and removed the overhanging DNA region with S1 nuclease. In the other, we hybridized 40S RNA to a 221 bp fragment of ribosomal DNA. The nucleotides encoding the 5' end of the 40S RNA were located more precisely by two methods. In one, we hybridized 40S RNA to the 221 bp DNA fragment and removed the overhanging DNA region with S1 nuclease. In the other, we hybridized 40S RNA to a smaller DNA fragment and extended the recessed 3' terminus of the DNA using reverse transcriptase. The resultant DNA fragments were sized on sequencing gels. Both determinations map the 5' end of 40S RNA at the same site in the rDNA, about 2250 bp upstream from the Eco RI site in the 18S rRNA coding sequence. At this site we find a DNA sequence beginning AGGGGAAGAC.... which agrees with partial sequence data from the 5' end of polyphosphorylated and bulk 40S rRNA. Features of this region of the ribosomal DNA will be discussed in this paper. A 227 nucleotide region surrounding the initiation site was also sequenced from an independently derived clone and found to differ in only one nucleotide. In addition, a sequence is found about 1100 nucleotides upstream from the 5' end of the gene that has 90% homology to the sequence from nucleotides minus 125 to +4 in the initiation region. At the termination region, X. laevis ribosomal DNA has a single recognition site for the restriction enzyme Hind III in each repeating unit. Using the S1 nuclease technique, the 3' termini of both the 40S precursor and mature 28S rRNA are seen to map within this recognition sequence. The sequence surrounding the Hind III site has striking homology to termination sites recognized by other RNA polymerase classes. Sequences with similar features are also found upstream from the initiation site.
...
PMID:The nucleotide sequence of the initiation and termination sites for ribosomal RNA transcription in X. laevis. 49 80

The secondary structure of HeLa 18S rRNA was investigated by a combination of chemical and enzymatic probing techniques. Using four chemical reagents (DMS*, kethoxal, DEPC and CMCT) which react specifically with unpaired bases and two nucleases (RNase T1 and cobra venom nuclease) which cleave the ribopolynucleotides at unpaired guanines and helical segments, we have analyzed the secondary structure of the 5' domain of 18S rRNA isolated from HeLa 40S ribosomal subunits. The sites at which chemical modifications and nuclease cleavages occurred were identified by primer extension using synthetic deoxyoligonucleotides and reverse transcriptase. These studies led to the deduction of an intra-RNA pairing pattern from the available secondary structure models based on comparative sequence analysis. Apart from the general canonical pairing we have identified noncanonical U-U, G-A, A-G, A-C, C-A and G-G pairing in HeLa 18S rRNA. The differential reactivity of bases to chemical reagents has enabled us to predict the possible configuration of these bases in some of the noncanonical pairing. The absence of chemical reactivities and cobra venom nuclease sensitivity in the terminal loops of helices 6 and 12 indicate a tertiary interaction unique to HeLa 18S rRNA. We have confirmed the existence of the complex tertiary folding recently proposed (Gutell and Woese 1990 Proc. Natl. Acad. Sci. 87, 663-667) for the universally conserved helix 19 in HeLa 18S rRNA. The complementarity of chemical modifications and enzymatic cleavages provided experimental evidence for the proposal of a model structure for the 655 nucleotides of the 5' domain of HeLa 18S rRNA.
...
PMID:Structural analysis of the 5' domain of the HeLa 18S ribosomal RNA by chemical and enzymatic probing. 226 64

Five of five human teratocarcinomas cultured in vitro could be induced to synthesize retrovirus-like particles, albeit in extremely low amounts. The accumulation and purification of the human teratocarcinoma-derived retrovirus (HTDV) from one of these cell lines allowed characterization of its genomic material as high mol. wt. (60S) RNA. Partial purification of HTDV-associated RNA-dependent DNA polymerase (reverse transcriptase) has also been achieved. HTDVs are easily distinguishable from the exogenous human T-lymphotropic viruses and human immunodeficiency viruses by morphological, biological and immunological criteria.
...
PMID:Genome analysis and reverse transcriptase activity of human teratocarcinoma-derived retroviruses. 244 5

The location of unpaired adenine residues within the secondary structure of rabbit 18S ribosomal RNA was determined by chemical probing. Naked 18S rRNA was first prepared by digestion of purified 40S subunits with matrix-bound proteinase K in sodium dodecyl sulfate, thereby omitting the use of nucleic acid denaturants. Adenines within naked 18S rRNA were chemically probed by using either diethyl pyrocarbonate or dimethyl sulfate, which specifically react with unpaired nucleotides [Peattie, D. A., & Gilbert, W. (1980) Proc. Natl. Acad. Sci. U.S.A. 77, 4679-4682]. Adenine modification sites were identified by polyacrylamide sequencing gel electrophoresis either upon aniline-induced strand scission of 32P-end-labeled intact and fragmented rRNA or by primer extension using sequence-specific DNA oligomers with reverse transcriptase. The data indicate good agreement between the general pattern of adenine reactivity and the location of unpaired regions in 18S rRNA determined by comparative sequence analysis [Chan, Y.-L., Gutell, R., Noller, H. F., & Wool, I. G. (1984) J. Biol. Chem. 259, 224-230]. The overall reactivity of adenine residues toward single-strand-specific chemical probes was, also, similar for both rabbit and Escherichia coli small rRNA. The number of strongly reactive adenines appearing within phylogenetically determined helical segments, however, was greater in rabbit 18S rRNA than for E. coli 16S rRNA. Some of these adenines were found clustered in specific helices. Such differences suggest a greater irregularity of many of the helical elements within mammalian 18S rRNA, as compared with prokaryotic 16S rRNA. These helical irregularities could be important for protein association and also may represent biologically relevant flexible regions of the molecule.
...
PMID:Chemical probing of adenine residues within the secondary structure of rabbit 18S ribosomal RNA. 334 49

The simultaneous assay of the reverse transcriptase and 60S to 70S RNA of oncogenic RNA viruses is described. The virus can be detected at low concentrations in biological fluids containing enzymatically active cell fragments or other contaminants. The fact that two features diagnostic of the oncornaviruses are used in the tests increases the certainty with which a positive outcome can be interpreted.
...
PMID:Simultaneous detection of reverse transcriptase and high molecular weight RNA unique to oncogenic RNA viruses. 433 Apr 70

Reaction conditions for Rous sarcoma virus ribonucleic acid (RNA)-instructed deoxyribonucleic acid (DNA) polymerase activity are described whereby the viral RNA is relatively protected from endogenous or added nuclease activity. Three analyses of reaction product nucleic acids ((3)H-RNA, (32)P-DNA) were compared, namely, gel electrophoresis, Cs(2)SO(4) gradient centrifugation, and hydroxyapatite column chromatography. It was found that hydroxyapatite analysis could be misleading unless the state of the template RNA was monitored concomitantly with the DNA analysis. Gel electrophoresis and Cs(2)SO(4) gradient centrifugation gave comparable results. It was concluded that analyses of the product of reverse transcriptase reactions should not only refer to the template RNA and product DNA species, but also be performed with virus or viral RNA which do not have or obtain nicks in the 60S RNA. Otherwise, interpretation of the results would have the ambiguity of potential artifacts caused by those degraded RNA molecules.
...
PMID:Deoxyribonucleic acid polymerase of Rous sarcoma virus: reaction conditions and analysis of the reaction product nucleic acids. 433 43

Particles from human milk contain a reverse transcriptase and a high-molecular-weight (60S to 70S) RNA that serves as a template. These particles have two features diagnostic of known RNA tumor viruses.
...
PMID:Detection of high-molecular-weight RNA in particles from human milk. 433 22

Affinity labeling of human placental 80S ribosomes with mRNA analogs of up to 12 uridyl residues, i.e. alkylating derivatives of oligouridylates bearing either 4-(N-2-chloroethyl-N-methylamino)benzylmethylphosphamide group at the 5'-termini or 2',3'-O-[4-(N-2-chloroethyl-N-methylamino)]benzylidene residue attached to the 3'-termini, in the presence of cognate Phe-tRNA(Phe) has been investigated. All the mRNA analogs modified only the 40S subunit. The fraction of 18S rRNA modified by the mRNA analogs with the alkylating group at the 5'-end decreased dramatically with extension of the reagent oligouridylate moiety. Nucleotides of 18S rRNA alkylated with the mRNA analogs were determined using a reverse transcription technique. For the mRNA analogs with the alkylating groups at the 3'-termini, G1702 and G1763/G1764 were identified as the cross-linking sites. The intensities of the bands corresponding to reverse transcriptase stops depended on the length of the reagent oligouridylate moieties. Cross-linking sites of the mRNA analogs with the alkylating group at the 5'-termini on 18S rRNA were A1023, C1026, C1057 and A1058 for the (pU)3 and (pU)4 derivatives and a single nucleotide C1057 for the (pU)6 one. Ribosomal protein S26 was found as the main target of modification with the same derivatives of (pU)6 and (pU)12.
...
PMID:Arrangement of mRNA at the decoding site of human ribosomes. 18S rRNA nucleotides and ribosomal proteins cross-linked to oligouridylate derivatives with alkylating groups at either the 3' or the 5' termini. 800 89

Ribosomal RNA genes are present near the end of the short arm and, to a lesser extent, near the centromere of the B chromosomes of some populations of Brachycome dichromosomatica. The internal transcribed spacer (ITS2) was amplified by PCR from total leaf DNA using primers within the conserved regions encoding the 5.8S and 25S stable rRNA species. Comparison of PCR amplified ITS2 sequences from several individual plants without B chromosomes with corresponding sequences derived from microdissected B chromosomes revealed two consistent differences between the rDNA of A and B chromosomes. One of these differences produced an SfcI restriction site that was present only in the ITS2 of the B-chromosome rDNA. Amplification by PCR of ITS2 from total genomic DNA from plants with and without B chromosomes showed an additive relationship between the amount of PCR product containing the SfcI site and the number of B chromosomes present. Quantitative analysis indicated that the proportion of total nuclear rDNA present on a single B chromosome varied between 2 and 4% in different A chromosome backgrounds. Similar experiments, with appropriate positive and negative controls, using reverse transcriptase PCR of the equivalent region within the 40S precursor rRNA, suggested that the B-chromosome rDNA was not transcribed. Similarly, PCR of reverse transcribed total RNA from plants containing B chromosomes using primers specific for the B chromosome ITS2 was unable to detect a transcript from the B chromosome.
...
PMID:Ribosomal RNA genes specific to the B chromosomes in Brachycome dichromosomatica are not transcribed in leaf tissue. 935 46

Conditions are described for using a primer extension inhibition (toeprinting) assay to study the initiation step of protein synthesis in rabbit reticulocyte lysates. These studies revealed that chloramphenicol acetyltransferase mRNA, which is widely used as a reporter, forms unusually labile initiation complexes. This and other unexpected problems were solved by adjustments in pH and temperature during the reverse transcriptase step. Complications that may occur during the ribosome binding step were also examined, including the possibility of rapid mRNA degradation. The suitability of inhibitors commonly used to block the elongation phase of translation was studied. The refined toeprinting assay was used to confirm context-dependent selection of the AUG start codon. Absence of the m7G cap did not subvert the process wherein initiation is restricted to the first AUG codon. The fidelity of initiation was impaired, however, when NaF was introduced during the ribosome binding step. In a preliminary assessment of the processivity of scanning, no dissociation of 40S ribosomal subunits was detected as the distance from the cap to the AUG codon was increased to nearly 300 bases. With an mRNA that contains a pseudoknot upstream from the AUG codon, the toeprinting assay revealed 40S ribosomal subunits trapped behind the base paired structure. Thus the assay is usable for mapping some intermediates as well as for detecting conventional 80S initiation complexes.
...
PMID:Primer extension analysis of eukaryotic ribosome-mRNA complexes. 977 44


<< Previous 1 2 3 Next >>

---