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:2.7.7.48 (transcriptase)
9,479 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

As an initial step toward investigating the roles of poliovirus proteins in viral RNA replication, a baculovirus expression system was used to produce poliovirus proteins from the P3 region. Spodoptera frugiperda (Sf9) cells were infected with a recombinant baculovirus, vETL-PoV3A*BCD, which contains cDNA coding for poliovirus proteins 3D, 3C, 3B, and a portion of 3A protein sequence. Immunofluorescence microscopy revealed that the majority of 3D (polymerase) was in the cytoplasm of recombinant baculovirus-infected Sf9 cells. In the same cells, the 3C (protease) and 3B (VPg) proteins appeared to be located in distinct subcellular regions, possibly membrane structures, suggesting that the expressed polyprotein was cleaved to generate mature proteins. Processing of the polypeptide was confirmed by immunoblot analysis which demonstrated that 3Cpro sequences were active in cleavage of the polyproteins 3A*BCD and 3CD. Over 95% of the 3D sequences accumulated in the form of mature 3Dpol, with only low levels of 3CD remaining. The majority of 3Dpol remained in the supernatant after low speed centrifugation of sonicated cells. The 3Dpol had RNA-dependent RNA polymerase activity as measured by elongation of an oligo(U) primer using a poly(A) template. The protein 3CDpro was active in cleaving P1 protein. The yield and activities of the poliovirus proteins expressed will facilitate future biochemical studies.
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PMID:Expression and characterization of poliovirus proteins 3BVPg, 3Cpro, and 3Dpol in recombinant baculovirus-infected Spodoptera frugiperda cells. 165 15

Reverse transcriptase has been purified from feline immunodeficiency virus (FIV) by DEAE-cellulose and phosphocellulose chromatography. The purified enzyme consists of a single protein with a Mr of 67,000. When proteolysis is not minimized during purification, a fragment of Mr 54,000 is also observed. This is similar to the reverse transcriptase from human immunodeficiency virus type 1 (HIV), which consists of a polypeptide of Mr 66,000; when proteolysis is not minimized during purification, a fragment of Mr 51,000 is also observed. In direct comparisons, the FIV reverse transcriptase is very similar to the HIV reverse transcriptase in template specificity and requirements for Mg2+. In contrast to these similarities, the FIV and HIV reverse transcriptases are substantially different in primary sequence, as determined by peptide mapping.
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PMID:Characterization of reverse transcriptase from feline immunodeficiency virus. 169 Jul 35

The nucleotide sequence of the large (L) genomic RNA segment of Seoul 80-39 virus was determined from overlapping cDNA clones. The virion L RNA segment is 6530 nucleotides long. The 3' and 5' terminal sequences are inversely complementary for 15 bases. The viral complementary-sense RNA contains a single open reading frame from an AUG codon at nucleotide position 37-39 to a UAA stop codon at nucleotide position 6490-6492. This ORF could encode a polypeptide of 2151 amino acids (246,662 kDa) which likely corresponds to the L protein detected in purified viral particles (Elliott et al., 1984) and is assumed to be an RNA-dependent RNA polymerase molecule (Schmaljohn and Dalrymple, 1983). Comparison of the L protein of the Seoul 80-39 virus with the polymerase proteins encoded by other negative-stranded RNA viruses revealed 44% similarity only with the part of the Bunyamwera virus L protein (Elliott, 1989) and a very weak homology with the PB1 protein of influenza virus.
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PMID:Nucleotide sequence and coding capacity of the large (L) genomic RNA segment of Seoul 80-39 virus, a member of the hantavirus genus. 184 Jul 13

Computer-assisted analysis of the putative polypeptide products encoded by the two open reading frames present in a large virus-like double-stranded RNA, L-dsRNA, associated with hypovirulence of the chestnut blight fungus, Cryphonectria parasitica, revealed five distinct domains with significant sequence similarity to previously described conserved domains within plant potyvirus-encoded polyproteins. These included the putative RNA-dependent RNA polymerase, RNA helicase, two papain-like cysteine proteases related to the potyvirus helper-component protease, and a cysteine-rich domain of unknown function similar to the N-terminal portion of the potyvirus helper-component protein. Phylogenetic trees derived from the alignment of the polymerase domains of L-dsRNA, a subset of positive-stranded RNA viruses, and double-stranded RNA viruses, using three independent algorithms, suggested that the hypovirulence-associated dsRNA and potyvirus genomes share a common ancestry. However, comparison of the organization of the conserved domains within the encoded polyproteins of the respective viruses indicated that the proposed subsequent evolution involved extensive genome rearrangement.
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PMID:Evidence for common ancestry of a chestnut blight hypovirulence-associated double-stranded RNA and a group of positive-strand RNA plant viruses. 196 31

The 8534 nucleotide sequence of the genome of the carlavirus, potato virus M (PVM), has been determined. The sequence contains six large open reading frames (ORFs) and non-coding regions consisting of 75 nucleotides at the 5' end, 70 nucleotides followed by a poly(A) tail at the 3' end and 38 and 21 nucleotides between three large blocks of coding sequences. The ORF beginning at the first initiation codon at nucleotide 76 encodes a polypeptide of 223K which, according to its primary sequence analysis, seems to be a virus RNA replicase. The next coding block consists of three ORFs encoding polypeptides of 25K, 12K and 7K. The third block consists of two ORFs encoding polypeptides of 34K (PVM coat protein) and 11K. The 11K polypeptide contains a pattern resembling the consensus for a metal-binding nucleic acid-binding 'finger'.
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PMID:The genome organization of potato virus M RNA. 199 70

The complete nucleotide sequence of the second largest RNA segment of Dhori/India/1313/61 virus was determined and the deduced amino acid sequence was compared with the polymerase (P) proteins of influenza A, B, and C viruses. RNA segment 2 (2224 nucleotides) of Dhori virus contains a single long open reading frame that can encode a 716-amino acid polypeptide (81.3 kDa). The predicted polypeptide shares between 27 and 31% sequence identities with the PB1 polypeptides of influenza A, B, and C viruses. Among the regions most highly conserved are the sequences around the Asp-Asp motif common to many RNA polymerases. In spite of the high level of sequence identity between the Dhori RNA segment 2 gene product and the influenza A, B, and C virus PB1 proteins the amino acid composition of the Dhori protein indicates an acidic charge feature at pH 7.0 in contrast to the basic nature of the PB1 proteins of the influenza viruses. We suggest that the Dhori PB1-like protein be designated the P alpha protein of this virus.
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PMID:Evolutionary relatedness of the predicted gene product of RNA segment 2 of the tick-borne Dhori virus and the PB1 polymerase gene of influenza viruses. 202 57

A region of the feline calicivirus (FCV) genome was sequenced which encodes polypeptides that are similar by amino acid alignment analysis to several picornavirus polypeptides. These polypeptides include the 2C polypeptide, the 3C cysteine protease and the 3D RNA-dependent RNA polymerase. The 2C-like region of the FCV genome encodes a GxxGxGKT nucleotide binding motif as well as amino acids which have been shown to be conserved in the picornavirus 2C polypeptides. The FCV RNA-dependent RNA polymerase also shows regions of similarity with picornavirus RNA polymerase sequences including the GDD sequence which is thought to be in or near the active site of the polymerase. The FCV cysteine protease-like sequences have the lowest degree of similarity with picornavirus cysteine proteases of the three regions aligned. However, the cysteine and histidine residues thought to be in the active site of the protease are present and are surrounded by amino acids conserved in the picornavirus cysteine proteases. The order of the polypeptides encoded in the FCV genome is the same as in the picornaviruses with the RNA-dependent RNA polymerase being located at the C-terminus of the FCV polyprotein. However, there is an approximately 40,000 dalton region between the FCV 2C- and the cysteine protease-like polypeptides which has no similarity to any known picornavirus protein. A striking difference between the organization of these sequences in FCV and the picornaviruses is that in the FCV genome, these non-structural polypeptides are encoded near the 5' end of the genomic RNA. Termination of the reading frame encoding these polypeptides occurs approximately 2400 bases from the 3' end of the genomic RNA as compared to 71 bases in the poliovirus genomic RNA.
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PMID:Nucleotide sequence of a region of the feline calicivirus genome which encodes picornavirus-like RNA-dependent RNA polymerase, cysteine protease and 2C polypeptides. 207 82

Nonstructural proteins of plum pox potyvirus were partially purified following a procedure described for the isolation of tobacco etch virus nuclear inclusion proteins. Plum pox virus proteins with electrophoretic mobilities corresponding to 49, 59 and 68 kDa reacted with antibodies against the 49 kDa and 54 kDa components of the nuclear inclusions and the 70 kDa component of the cylindrical inclusions of tobacco etch virus, respectively. Further purification by size exclusion high performance liquid chromatography or SDS-polyacrylamide gel electrophoresis, and amino terminal amino acid sequencing permitted the location in the plum pox virus polyprotein of the cleavage sites from which the 49 kDa (NIa-type, protease), 59 kDa (NIb-type, putative RNA replicase), and 68 kDa (CI-type) proteins originate. A 110 kDa protein which copurified with the plum pox virus inclusion proteins reacted with both anti-NIa and anti-NIb sera and had the same amino terminus as the plum pox virus 49 kDa protein, indicating that it is a non-processed 49-59 kDa polypeptide.
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PMID:Determination of polyprotein processing sites by amino terminal sequencing of nonstructural proteins encoded by plum pox potyvirus. 213 35

The double-stranded RNA (dsRNA) viruses of Saccharomyces cerevisiae consist of 4.5-kilobase-pair (kb) L species and 1.7- to 2.1-kb M species, both found in cytoplasmic viruslike particles (VLPs). The L species encode their own capsid protein, and one (LA) has been shown to encode a putative capsid-polymerase fusion protein (cap-pol) that presumably provides VLPs with their transcriptase and replicase functions. The M1 and M2 dsRNAs encode the K1 and K2 toxins and specific immunity mechanisms. Maintenance of M1 and M2 is dependent on the presence of LA, which provides capsid and cap-pol for M dsRNA maintenance. Although a number of different S. cerevisiae killers have been described, only K1 and K2 have been studied in any detail. Their secreted polypeptide toxins disrupt cytoplasmic membrane functions in sensitive yeast cells. K28, named for the wine S. cerevisiae strain 28, appears to be unique; its toxin is unusually stable and disrupts DNA synthesis in sensitive cells. We have now demonstrated that 4.5-kb L28 and 2.1-kb M28 dsRNAs can be isolated from strain 28 in typical VLPs, that these VLPs are sufficient to confer K28 toxin and immunity phenotypes on transfected spheroplasts, and that the immunity of the transfectants is distinct from that of either M1 or M2. In vitro transcripts from the M28 VLPs show no cross-hybridization to denatured M1 or M2 dsRNAs, while L28 is an LA species competent for maintenance of M1. K28, encoded by M28, is thus the third unique killer system in S. cerevisiae to be clearly defined. It is now amenable to genetic analysis in standard laboratory strains.
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PMID:K28, a unique double-stranded RNA killer virus of Saccharomyces cerevisiae. 220 3

A soluble RNA-dependent RNA polymerase was isolated from Nicotiana tabacum plants infected with cucumber mosaic virus (CMV), which has a genome of three positive-strand RNA components, 1, 2, and 3. The purified polymerase contained two virus-encoded polypeptides and one host polypeptide. Polymerase activity was completely dependent on addition of CMV RNA as template, and the products of reaction were single-stranded (ss) RNA and double-stranded (ds) RNA, corresponding to RNAs 1, 2, and 3, and a subgenomic RNA (RNA 4) derived from RNA 3. The ratio of ssRNA to dsRNA was about 5:1, and the ssRNA was shown to be predominantly the positive strand. This demonstrates the complete replication of a eukaryotic virus RNA in vitro by a template-dependent RNA polymerase.
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PMID:Complete replication of a eukaryotic virus RNA in vitro by a purified RNA-dependent RNA polymerase. 220 91


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