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)

A Trypanosoma brucei repeated DNA element (TRS-1, for trypanosome repeated sequence), which seems transposable, may encode a 1651 amino acid polypeptide showing homology with reverse transcriptase. This polypeptide would also carry a DNA-binding domain, as suggested by the presence of five DNA-binding "fingers" homologous to those of the transcription factor TFIIIA of Xenopus laevis and retroviral DNA-binding proteins.
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PMID:DNA-binding fingers encoded by a trypanosome retroposon. 244 94

The causative agent of AIDS the human immunodeficiency virus (HIV) encodes as part of its pol gene a reverse transcriptase (RT) which has a key role in the replication of the virus and thus constitutes an ideal target for antiviral chemotherapy. The purified HIV RT from virus particles consists of two related polypeptides of 66 and 51 kd mol. wt and similar polypeptides are found on expression of the complete HIV pol gene using prokaryotic systems. Here we describe the expression of the 66-kd protein in Escherichia coli and demonstrate that this polypeptide alone has authentic RT activity. Thus, a central HIV pol gene segment encodes and is sufficient for high levels of RT activity. The RT has been purified from E. coli extracts using a purification procedure involving two chromotography steps resulting in an enzyme preparation near homogeneity. Deletion of the C-terminal region of the RT thought to encode the RNase H domain resulted in loss of polymerase activity.
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PMID:AIDS virus reverse transcriptase defined by high level expression in Escherichia coli. 244 66

Retroviral reverse transcriptase possesses DNA polymerase and ribonuclease H (RNase H) activity within a single polypeptide. Chemical or proteolytic treatment of reverse transcriptase has been used in the past to produce enzyme that is missing DNA polymerase activity and retains RNase H activity. It has not been possible to obtain reverse transcriptase that lacks RNase H but retains DNA polymerase activity. We have constructed a novel deletion derivative of the cloned Moloney murine leukemia virus (M-MLV) reverse transcriptase gene, expressed the gene in E. coli, and purified the protein to near homogeneity. The purified enzyme has a fully active DNA polymerase, but has no detectable RNase H activity. These results are consistent with, but do not prove, the conclusion that the DNA polymerase and RNase H activities of M-MLV reverse transcriptase reside within separate structural domains.
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PMID:Isolation of cloned Moloney murine leukemia virus reverse transcriptase lacking ribonuclease H activity. 244 47

Human immune deficiency virus (HIV) replicates by conversion of the RNA genome into the double-stranded DNA provirus. The reverse transcriptase is not the only enzymatic function crucial in DNA-provirus synthesis. A viral-coded RNase H activity which specifically degrades RNA in RNA-DNA hybrids has been shown to be essential as well. Here we demonstrate that the HIV-reverse transcriptase which consists of a two-polypeptide complex, p66 and p51, copurifies with an RNase H activity which exhibits properties of a processive exonuclease. Only the p66 molecule, not p51, is active as polymerase as evidenced by activated gel analysis. p66 exhibits RNase H activity when precipitated as immune complex by a monoclonal antibody raised against a bacterially expressed carboxy-terminal portion of p66. The monoclonal antibody which does not interfere with enzyme activity also precipitates a second population of molecules with RNase H activity which is of low mol. wt, p15. This RNase H appears therefore to be derived from the carboxy terminus of p66 during processing to the p51 polypeptide. It exhibits low template-binding ability and is of a non-processing mode of action which may be due to the absence of the reverse transcriptase domain. These results lend experimental support to the hypothesis that the RNase H gene maps at the carboxy terminus of the reverse transcriptase. Since both RNase H populations are virus-coded they may be essential for retrovirus replication in general and useful targets for chemotherapeutic agents.
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PMID:Identification and characterization of HIV-specific RNase H by monoclonal antibody. 245 83

We have cloned the entire pol gene of human immunodeficiency virus type 2 into a high-level Escherichia coli expression system. Induction of cultures containing the recombinant plasmid, p2RTL1, leads to rapid accumulation of polypeptides of 66, 54, and 34 kilodaltons. We have designated the larger polypeptides reverse transcriptase, and we have designated the smaller polypeptide endonuclease. Purification of reverse transcriptase via ion-exchange and affinity chromatography yields the 66-kilodalton polypeptide, with which reverse transcriptase activity is associated. Purified enzyme furthermore displays a higher apparent molecular weight than its counterpart from human immunodeficiency virus type 1.
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PMID:A single 66-kilodalton polypeptide processed from the human immunodeficiency virus type 2 pol polyprotein in Escherichia coli displays reverse transcriptase activity. 245 82

The reverse transcriptase of HIV-1 (AIDS virus) is characterized by the presence of two highly immunogenic proteins of 66 and 51 kD known to be enzymatically active as a complex p66/51. Using an activity gel procedure that allows identification of catalytic polypeptides in situ after PAGE in denaturing conditions, we visualized two major active bands of 66 and 51 kD of reverse transcriptase from highly purified preparations of HIV-1. We show that both p66 and p51 are enzymatically active. An additional active band was also associated with a 165 kD polypeptide, representing about 2-4% of total activity and possibly corresponding to the putative gag-pol precursor. In H9-infected cells the 66 kD active band became visible 70 hours after infection. These studies show that the two major forms of reverse transcriptase (66 and 51 kD) of HIV-1 are independently active and that a higher Mr form of 165 kD is also enzymatically active.
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PMID:Enzymatically active forms of reverse transcriptase of the human immunodeficiency virus. 246 25

From the yeast, Saccharomyces cerevisiae, three proteins exhibiting ribonuclease H activity were isolated. These proteins differ in molecular weights and enzymatic properties. The two smaller ones, RNAase H(55) and RNAase H(42) are immunologically and structurally related to each other. Neither reacts with antibodies against the largest one, RNAase H(70). Highly purified preparations of RNAase H(70) contain two polypeptides (Mr 70,000 and 160,000) and display reverse transcriptase activity. Deletion of part of the gene for the 160 kDa polypeptide results in mutants possessing about twice the amount of DNA as do wild-type cells. DNA polymerase stimulating activity resides in the 70,000 polypeptide. The processivity of yeast DNA polymerase A(I) does not change in presence of that protein. Possible functions of RNAases H are discussed.
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PMID:Three ribonucleases H and a reverse transcriptase from the yeast, Saccharomyces cerevisiae. 246 14

Myxobacteria have been shown to produce a peculiar RNA-DNA complex called msDNA, in which a single-stranded DNA is branched out from a RNA molecule (msdRNA) by a 2',5' phosphodiester linkage. It has been predicted that reverse transcriptase is required for msDNA biosynthesis. We identified a gene for reverse transcriptase in M. xanthus in the region that has been demonstrated to code for a cis- or transacting element for msDNA synthesis. This gene is located immediately downstream of the msdRNA coding region, and codes for a polypeptide of 485 amino acid residues. The polypeptide shows sequence similarity with retroviral reverse transcriptases. This fact, together with the mode of msDNA synthesis, suggests a possible relationship between retroviruses and the msDNA system. The analysis of the gene and the distribution of the msDNA system in independent isolates of M. xanthus indicate that the element is as old as other essential genes in M. xanthus and that it was not recently acquired into the genome.
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PMID:Reverse transcriptase associated with the biosynthesis of the branched RNA-linked msDNA in Myxococcus xanthus. 246 92

Bacterially expressed recombinant HIV-1 reverse transcriptase is active as both a homodimer of Mr 66,000 subunits and a heterodimer of Mr 66,000 and 51,000 subunits. The heterodimer is formed by cleavage of a C-terminal fragment from one Mr 66,000 polypeptide, which occurs during purification and crystallization of reverse transcriptase. Thus, crystals obtained from purified Mr 66,000 polypeptide preparations consisted of an apparently equimolar mixture of Mr 66,000 and 51,000 polypeptides, which were apparently analogous to the Mr 66,000 and 51,000 polypeptides detected in HIV-infected cells and in virions. Limited proteolysis of the homodimer with alpha-chymotrypsin also resulted in cleavage to a stable Mr 66,000/51,000 mixture, and proteolysis with trypsin resulted in the transient formation of some Mr 51,000 polypeptide. These results are consistent with the reverse transcriptase molecule having a protease-sensitive linker region following a structured domain of Mr 51,000. Further digestion with trypsin resulted in cleavage of the Mr 51,000 polypeptide after residue 223, yielding peptides of apparent Mr 29,000 and 30,000. A minor peptide of Mr 40,000 was also produced by cleavage of the Mr 66,000 polypeptide after residue 223. About half the original Mr 66,000 polypeptides remained resistant to proteolysis and existed in complex with the above peptides in solution. During both chymotrypsin and trypsin digestion there was an increase in the reverse transcriptase activity caused by a doubling of Vmax with little change in Km for dTTP.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:HIV-1 reverse transcriptase: crystallization and analysis of domain structure by limited proteolysis. 246 81

Open reading frame (ORF) V of cauliflower mosaic virus (CaMV), the candidate for the reverse transcriptase gene, has been expressed in E. coli under control of the PR promoter of bacteriophage lambda either as an N-terminal polypeptide fused to beta-galactosidase or as the total ORF V without fusion. Antibodies against these proteins were used to analyze extracts from CaMV-infected plants by immunoblotting. ORF V-specific polypeptides of 80, 62, 58, 22, and 18 kD apparent molecular weights were detected, with the largest species corresponding to the full length translation product. The 62 and 22 kD species could be assigned to the N-terminus and the remaining two species to the C-terminus of the ORF.
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PMID:Translation products of cauliflower mosaic virus ORF V, the coding region corresponding to the retrovirus pol gene. 246 52


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