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 mutant of the Bryan high-titer strain of Rous sarcoma virus defective in reverse transcriptase is known as type alpha (BH-RSV alpha). BH-RSV alpha virion particles do not contain any polymerase-related proteins but they direct the synthesis of a normal sized Pr180 gag-pol polyprotein precursor in infected cells. Using a bioassay for polymerase gene function that is based on the requirement of viral replication for transformation of transfected chicken cells, we have localized the defect to the 2.5 kb EcoRI-KpnI DNA fragment containing more than 90% of the polymerase gene by comparison with the corresponding DNA fragment from the wild-type polymerase-positive BH-RSV, called type beta. In vitro recombination experiments with the polymerase gene of Schmidt-Ruppin RSV allowed us to map the defect to the 0.86 kb XbaI-BglII DNA fragment of the BH-RSV alpha polymerase. DNA sequence analysis of the entire polymerase gene of BH-RSV alpha and beta has revealed one point mutation that maps within that XbaI-BglII fragment and substitutes leucine in BH-RSV alpha for glutamine in the wild-type BH-RSV beta.
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PMID:Polymerase-defective mutant of the Bryan high-titer strain of Rous sarcoma virus. 242 Dec 48

The nucleotide sequence of the internal region of a Drosophila retrotransposon. 412, was determined. The genome of 412 was found to consist of two long open-reading frames (ORFs 1 and 2), an unusually long putative leader region and long terminal repeats (LTRs). As with 17.6, 297 and gypsy, ORFs 1 and 2 slightly overlap each other and are out of phase by +2. ORF2 includes the nucleotide sequences coding for the putative protease, reverse transcriptase and integrase, and is similar in entire organization to the pol gene of Moloney murine leukaemia virus. In spite of the difference in insertion specificity, integrase, an enzyme presumably responsible for insertion, was found to be similar in amino acid sequence to the counterparts of 17.6, 297 and gypsy. There is no ORF in 412 which corresponds to retroviral env or ORF3s of 17.6 and 297. Analysis of 412 transcripts suggested that 412 LTR is composed of U3, R and U5. The gene for a potential primer tRNA for putative reverse transcription of 412 was also surveyed and the 3'-terminal 15 nucleotides of a putative arginine tRNA were found to be exactly complementary to the putative primer-binding site of 412.
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PMID:Nucleotide sequence characterization of a Drosophila retrotransposon, 412. 242 8

Portions of the pol gene of Moloney murine leukemia virus (MuLV) were expressed as fusion proteins in Escherichia coli, and the purified proteins were used to elicit antibodies in Escherichia coli, and the purified proteins were used to elicit antibodies in rabbits. The sera were used to examine the mature pol gene products contained in virion particles and identified the reverse transcriptase and a second protein, P46pol, encoded by the 3' portion of the gene. The P46 protein was not phosphorylated and was present at the same molar abundance as the reverse transcriptase. The sera were also used to detect the Pr200gag-pol intracellular precursor protein and to analyze its processing to the mature forms. The proteins formed by several Moloney MuLV mutants were analyzed. Further tests revealed cross-reactivity with Friend MuLV and feline leukemia virus proteins, but not with avian retrovirus proteins.
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PMID:Analysis of retroviral pol gene products with antisera raised against fusion proteins produced in Escherichia coli. 242 63

The pol gene from a biologically active clone of the human T-cell lymphotropic virus type III provirus was inserted into a bacterial expression vector. The resulting gene fusion induced the formation of active reverse transcriptase that could be readily detected in extracts of bacterial cells. The activity exhibited the template and divalent cation requirements of the authentic enzyme. These constructs will be useful for safe and rapid analysis of potential inhibitors of this important enzyme.
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PMID:Expression of reverse transcriptase activity of human T-lymphotropic virus type III (HTLV-III/LAV) in Escherichia coli. 242 71

Yeast cells contain a protein of molecular size 70 kDa that possesses RNase H activity. A polyclonal antibody against it reacts in addition with proteins of molecular sizes 160 kDa from yeast extracts. All these immunologically related proteins exhibit reverse transcriptase activity and in this respect they resemble the products of retroviral pol genes, relatives of which reside in Ty elements and mitochondrial introns of yeast. Experimental evidence, however, indicates that the protein described here that combines RNase H and reverse transcriptase activity is not coded for by a known element of the retrotransposon family. It may originate from a cellular gene distantly related to retrotransposon sequences.
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PMID:Ribonuclease H(70) from Saccharomyces cerevisiae possesses cryptic reverse transcriptase activity. 242 7

We have cloned and expressed in Escherichia coli a section of the Moloney murine leukemia virus (Mo-MLV) pol gene which includes the entire coding region of mature reverse transcriptase (RT) plus 284 additional base pairs 3' to the coding region (Kotewicz et al., 1985). To prepare cloned Mo-MLV RT as close as possible to authentic RT in structure and activity, the universal terminator sequence GC(TTAA)3GC was introduced at a number of positions inside and outside the RT coding region within 200 nucleotides of its 3' end. The level of RT activity expressed from these constructs varied sevenfold. This variation was found to be directly related to the stability of the RT protein products in the E. coli K-12 strain K802; half-lives varied from 2 to 35 min. The stability of most of the RT proteins was not increased in E. coli K802 lon- cells, with the exception of two, whose half-lives were increased by a factor of two.
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PMID:Influence on stability in Escherichia coli of the carboxy-terminal structure of cloned Moloney murine leukemia virus reverse transcriptase. 242 93

The organization of the murine leukemia virus (MuLV) pol gene was investigated by expressing molecular clones containing AKR MuLV reverse transcriptase or endonuclease or both gene segments in Escherichia coli and generating specific antisera against the expressed bacterial proteins. Reaction of these antisera with detergent-disrupted virus precipitated an 80-kilodalton (kDa) protein, the MuLV reverse transcriptase, and a 46-kDa protein which we believe is the viral endonuclease. A third (50-kDa) protein, related to reverse transcriptase, was also precipitated. Bacterial extracts of clones expressing reverse transcriptase and endonuclease sequences competed with the viral 80- and 46-kDa proteins, respectively. These results demonstrate that the antisera are specific for viral reverse transcriptase and endonuclease. Immunoprecipitation of AKR MuLV with antisera prepared against a bacterial protein containing only endonuclease sequences led to the observation that reverse transcriptase and endonuclease can be associated as a complex involving a disulfide bond(s).
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PMID:Murine leukemia virus pol gene products: analysis with antisera generated against reverse transcriptase and endonuclease fusion proteins expressed in Escherichia coli. 242 47

A computer analysis of the amino acid sequences from the putative gene products of retroviral pol genes has revealed a 150-residue segment that is homologous with the ribonuclease H of Escherichia coli. The segment occurs at the carboxyl terminus of the region assigned to the 90-kDa reverse transcriptase polypeptide. In contrast, a section nearer the amino terminus of this sequence can be aligned with nonretroviral polymerases. The order of activities in the pol gene is thus: polymerase-ribonuclease-endonuclease. On another note, all retroviral endonuclease sequences contain a consensus zinc-binding "finger." This should not be confused with the well-known zinc requirement of reverse transcriptases.
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PMID:Computer analysis of retroviral pol genes: assignment of enzymatic functions to specific sequences and homologies with nonviral enzymes. 242 13

We have found human DNA to contain a number of sequences related to simian sarcoma associated virus (SSAV). One of these sequences was isolated from a human genomic library. The molecular clone, termed S71, contains regions homologous to SSAV gag and pol fragments and SSAV LTR. Furthermore, hybridization experiments and DNA sequencing revealed distinct homologies to the reverse transcriptase coding region of several other retroviruses including baboon endogenous virus (BaEV) and murine leukemia viruses (MuLV) as well as retrovirus-like elements. Some sequence homology was also found with the C-type retrovirus-related multicopy human clone 4-1. S71 is present in only one copy per human genome equivalent and exhibits an EcoRI restriction fragment length polymorphism.
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PMID:Isolation of an SSAV-related endogenous sequence from human DNA. 243 42

Three pol gene products have been identified in avian retroviral particles: the full-length 95-kilodalton (kDa) beta chain of reverse transcriptase and two proteolytic cleavage products of beta, a 63-kDa reverse transcriptase alpha chain derived from the amino terminus of beta and a 32-kDa (pp32) endonuclease from its carboxy terminus. By using molecularly cloned retroviral DNA and synthetic oligonucleotides to introduce initiator ATGs and codons corresponding to the authentic N termini, we constructed two bacterial-expression clones; one clone contains the entire pol gene, and the other contains the region encoding the pp32 domain. A 99-kDa protein was synthesized in Escherichia coli by the full-length clone, and a 36-kDa protein was synthesized by the endonuclease domain clone. The recombinant proteins exceeded the size of both the mature viral beta chain and the pp32, respectively, by approximately 4 kDa. These larger sizes, however, are consistent with predictions from the DNA sequence of the pol gene. Processing of the recombinant pol proteins was examined by using p15 protease purified from virus particles and antisera directed against synthetic peptides corresponding to three domains in pol. Proteolytic digestion of the 99-kDa product with p15 produced a 63-kDa protein that comigrated on polyacrylamide gels with the alpha chain of reverse transciptase and a 36-kDa fragment that comigrated with the endonuclease domain product. Further digestion of the 36-kDa protein yielded a 32-kDa protein that comigrated with viral pp32 endonuclease. Thus, we concluded that two p15-sensitive sites exist in pol. Cleavage at the previously identified site produces alpha, and cleavage at the newly discovered site removes approximately 4 kDa from the C terminus of the primary protein product. Since the 36-kDa protein was also detected in protein isolated from virus particles, it seems probable that processing at the C-terminal site is a normal step in the production of mature beta and pp32 endonuclease products.
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PMID:Proteolytic processing of avian sarcoma and leukosis viruses pol-endo recombinant proteins reveals another pol gene domain. 243 65


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