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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 genomic structure of the rat LINE (L1Rn) DNA element contains two overlapping open reading frames (ORFs) and apparently has a potential to code for a DNA/RNA-binding protein (in ORF1) and a reverse transcriptase (in ORF2). We have characterized a 1,630-bp L1Rn cDNA clone encompassing the overlapping ORFs and a 600-bp genomic fragment derived from a full-length L1Rn member and containing the beginning of ORF1. These DNAs were used to restore in part the ORF1-ORF2 organization of L1Rn after being cloned into the pSP65 vector under the control of SP6 polymerase promoter. To test whether L1Rn ORF1 and ORF2 are expressed as a fusion protein, a series of capped RNAs with progressive truncations containing one or both ORFs were prepared and translated in the rabbit reticulocyte lysate. Our analysis indicates that the expression of a putative reverse transcriptase-encoded L1Rn ORF2 in vitro is regulated by reinitiation or internal initiation of translation but not by ribosomal frameshifting.
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PMID:Translation of the rat LINE bicistronic RNAs in vitro involves ribosomal reinitiation instead of frameshifting. 138 Jun 49

The mobile element jockey is similar in structural organization and coding potential to the LINEs of various organisms. Current models of the mechanism of transposition involve reverse transcription of an RNA intermediate and utilization of element-encoded proteins. As it is demonstrated here, a 2.23 kb DNA fragment from the region of the jockey encoding the putative reverse transcriptase, was stably introduced into the expression system under inducible control of the Escherichia coli lac regulatory elements. We describe the expression of the 92 kDa protein and identify this polypeptide alone as authentic jockey reverse transcriptase based on some of its physical and enzymic properties. The jockey polymerase demonstrates RNA-directed and DNA-directed DNA polymerase activities, but lacks detectable RNase H, has a temperature optimum at 26 degrees C, requires Mg2+ or Mn2+ as a cofactor and is inactivated by sulfhydryl reagent. The enzyme prefers poly(rC) and poly(rA) as template and "activated" DNA is not effective. The results of this work suggest that the RNA-directed DNA polymerase coded by jockey elements may be involved in the transcription of the elements.
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PMID:[Cloning and expression in Escherichia coli of reverse transcriptase coded by the mobile genetic element jockey]. 138 Jun 45

The Mauriceville and Varkud mitochondrial plasmids of Neurospora are closely related, closed-circular DNAs (3.6 and 3.7 kilobases, respectively) that have characteristics of mtDNA introns and retroid elements. The plasmids contain a single long open reading frame (710 amino acids), whose amino-terminal half has structural similarity to reverse transcriptases. Using antibodies against synthetic peptides and trpE fusion proteins, we detected an 81-kDa protein encoded by this open reading frame in mitochondrial preparations from the plasmid-containing strains. This 81-kDa protein cosegregates with reverse transcriptase activity in sexual crosses and comigrates with reverse transcriptase activity in sodium dodecyl sulfate-polyacrylamide gels, where it can be assayed after renaturation of the protein. In glycerol gradients under nondenaturing conditions, the reverse transcriptase activity sediments at approximately 145 kDa, close to the value expected for a dimer of the 81-kDa protein. The 81-kDa protein represents most of the 710-amino acid open reading frame, but may be missing some amino acids at the amino terminus. The regions upstream and downstream of the putative reverse transcriptase domain lack sequences characteristic of gag, protease, RNase H, or integrase domains found in other retroid elements. The plasmid-encoded 81-kDa protein seems to be a novel type of reverse transcriptase that may provide insight into the evolution of these enzymes.
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PMID:Identification of the reverse transcriptase encoded by the Mauriceville and Varkud mitochondrial plasmids of Neurospora. 169 Nov 79

The mobile element jockey is similar in structural organization and coding potential to the LINEs of various organisms. It is transcribed at different stages of Drosophila ontogenesis. The Drosophila LINE family includes active transposable elements. Current models for the mechanism of transposition involve reverse transcription of an RNA intermediate and utilization of element-encoded proteins. As demonstrated here, a 2.23 kb DNA fragment from the region of jockey encoding the putative reverse transcriptase was stably introduced into an expression system under inducible control of the Escherichia coli lac regulatory elements. We describe the expression of the 92 kDa protein and identify this polypeptide alone as the authentic jockey reverse transcriptase based on some of its physical and enzymic properties. The jockey polymerase demonstrates RNA and DNA-directed DNA polymerase activities but lacks detectable RNase H, has a temperature optimum at 26 degrees C, requires Mg2+ or Mn2+ as a cofactor and is inactivated by sulphydryl reagent. The enzyme prefers poly(rC) and poly(rA) as template and 'activated' DNA is not effective.
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PMID:Authentic reverse transcriptase is coded by jockey, a mobile Drosophila element related to mammalian LINEs. 171 78

Long interspersed elements (LINEs) of the L1 family represent a major class of mammalian repetitive DNA and are present at copy numbers of between 10(4) and 10(5) elements per genome. Structural similarities between L1 elements and known retrotransposons have led to the suggestion that a subset of L1 elements may function as mobile genetic elements and have thus gained their prominent place in the mammalian genome. We describe a consensus mouse L1 element that was tagged with a heterologous intron and shown to transpose by way of an RNA intermediate when transfected into baby hamster kidney cells, formally establishing L1 elements as retrotransposons. When the putative reverse transcriptase-encoding region of this L1 element was deleted, the element still underwent retrotransposition in hamster cells, suggesting that reverse transcriptase activity can be supplied by an endogenous enzyme.
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PMID:Retrotransposition of a mouse L1 element. 171 87

A detailed investigation of the Drosophila melanogaster mobile dispersed repetitive element jockey was performed. This is similar in its structural organization and coding potential to the long interspersed elements (LINEs) of various organisms. A complete copy of jockey (approx. 5 kb) is terminated with an oligodeoxynucleotide (dA) sequence preceded by two long open reading frames (ORFs) overlapping with a frameshift-1. Judging by the sequence homologies, ORF1 codes for a nucleic-acid-binding protein, and ORF2 for a reverse transcriptase which is most similar in its sequence to putative reverse transcriptase of other LINEs. As demonstrated by sequencing two deleted jockey copies, they contain only a small part of ORF2; however, other regions, including the terminal sequences, are highly conservative. The existence of a large number of jockey copies with a deletion in the second frame may indicate that they can use reverse transcriptase in trans.
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PMID:The Drosophila mobile element jockey belongs to LINEs and contains coding sequences homologous to some retroviral proteins. 246 54

By means of comparative analysis of primary and secondary structures, and hydropathy plots of hepadnavirus P proteins new functional domains were revealed additionally to the polymerase domain which had been found earlier in these proteins. The C-terminal part of P proteins was revealed to be significantly similar to ribonuclease H of E. coli. The ribonuclease H functional domain is known to be an integral entity of retrovirus reverse transcriptase as a rule. Availability of this domain indicates once more the putative reverse transcriptase properties of the P products. The proteins of hepadnaviruses were compared to terminal proteins of picornaviruses, adenoviruses and bacteriophages. The data obtained suggested that a conservative N-terminal region of P proteins functions as protein primer for DNA synthesis in hepadnaviruses.
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PMID:Prediction of terminal protein and ribonuclease H domains in the gene P product of hepadnaviruses. 246 82

Hepatitis B virus (HBV), like retroviruses, replicates through reverse transcription. However, the identity and mechanism for the synthesis of HBV reverse transcriptase remain unknown. The open reading frame (ORF) for HBV putative reverse transcriptase (pol), as a consequence of overlapping with the whole ORF of envelope proteins (hepatitis B surface antigens), includes a hypervariable region at the N terminus. Thus, compared with retroviruses, it is unlikely that HBV reverse transcriptase is translated from complete pol ORF in the full-length pregenomic RNA. We have now detected in infected human livers a novel doubly spliced RNA in which one splicing event removed the hypervariable region of the pol gene but retained the conserved region homologous to retroviral reverse transcriptase. The other splicing event deleted the central region of hepatitis B core antigen and thus brought the protease domain which is important for maturation of reverse transcriptase close to that of pol. For this sequence organization, the spliced RNA as the possible template for the synthesis of HBV reverse transcriptase is discussed.
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PMID:Identification of a doubly spliced viral transcript joining the separated domains for putative protease and reverse transcriptase of hepatitis B virus. 247 67

Bovine leukemia virus protease was purified to homogeneity and assayed by using murine leukemia virus Pr65gag, a polyprotein precursor of the viral core structural proteins, as the substrate. A chemical analysis of the protease, including an amino acid composition and NH2- and COOH-terminal amino acid sequence analysis, revealed that it has an Mr of 14,000 and is encoded by a segment of the viral RNA located between the gag gene and the putative reverse transcriptase gene. As expected from the nucleotide sequence data (Rice et al., Virology 142:357-377, 1985), the reading frame for the protease is different from both the gag and reverse transcriptase reading frames. The 5' end of the protease open reading frame extends 38 codons upstream from the codon for the NH2-terminal residue of the mature viral protease and overlaps the gag open reading frame by 7 codons. The 3' end of the protease open reading frame extends 26 codons beyond the codon for the COOH-terminal residue of the mature protease and overlaps 8 codons of the reverse transcriptase open reading frame. Several lines of evidence, such as protein mapping of the gag polyprotein precursor, the characteristic structure of the mRNA, and promotion of the synthesis of a gag polyprotein precursor by lysine tRNA in vitro, suggest that the protease could be translated by frameshift suppression of the gag termination codon. In vitro synthesized bovine leukemia virus gag-related polyproteins were cleaved by the protease into fragments which were the same size as the known components of bovine leukemia virus, suggesting that the specificity of cleavage catalyzed in vitro by the purified protease is the same as the specificity of cleavage found in the virus.
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PMID:Bovine leukemia virus protease: purification, chemical analysis, and in vitro processing of gag precursor polyproteins. 300 29

VIPER was initially characterized as a 2326bp LTR-like retroelement associated to SIRE, a short interspersed repetitive element specific of Trypanosoma cruzi. It carried a single ORF that coded for a putative reverse transcriptase-RNAse H protein, suggesting that it could be a truncated copy of a longer retroelement. Herein we report the identification and characterization of a complete 4480bp long VIPER in the T. cruzi genome. The complete VIPER harbored three non-overlapped domains encoding for a GAG-like, a tyrosine recombinase and a reverse transcriptase-RNAse H proteins. VIPER elements were also found in the genomes of Trypanosoma brucei and Trypanosoma vivax, but not in Leishmania sp. On the basis of its reverse transcriptase phylogeny, VIPER was classified as an LTR retroelement. However, VIPER was structurally related to the tyrosine recombinase encoding retroelements, DIRS and Ngaro. Phylogenetic analysis showed that VIPER's tyrosine recombinase grouped with the transposases RCI1 of Escherichia coli and Ye24 and Ye72 of Haemophilus influenzae within a major branch of prokaryotic recombinases. Taken together, VIPER's structure, the nature of its tyrosine recombinase, the unique features of its reverse transcriptase catalytic consensus motif and the fact that it was found in Trypanosomes, an early branching eukaryote, suggest that VIPER may be the closest relative of the founder element of the tyrosine recombinase encoding retrotransposons known up to date. Our analysis revealed that tyrosine recombinase-encoding retroelements were originated as early in evolution as non-LTR retroelements and suggests that VIPER, Ngaro and DIRS elements may constitute a third group of retrotransposons, distinct from both LTR and non-LTR retroelements.
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PMID:The VIPER elements of trypanosomes constitute a novel group of tyrosine recombinase-enconding retrotransposons. 1629 62


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