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RNA-dependent RNA polymerase

About: RNA-dependent RNA polymerase is a research topic. Over the lifetime, 13904 publications have been published within this topic receiving 767954 citations. The topic is also known as: RdRp & RNA replicase.


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Journal ArticleDOI
TL;DR: Results indicate that the stem-loop structure called the dimerization initiation site is a cis element acting on both genomic RNA packaging and synthesis of proviral DNA.
Abstract: In retroviruses, the genomic RNA is in the form of a 60S-70S complex composed of two identical genome-length RNA molecules tightly associated through numerous interactions. A major interaction, called the dimer linkage structure, has been found near the RNA 5' end and is probably involved in the control of translation, packaging, and recombination during proviral DNA synthesis. Recently, a small sequence corresponding to a stem-loop structure located in the 5' leader of human immunodeficiency virus type 1 (HIV-1) RNA was found to be required for the initiation of HIV-1 RNA dimerization in vitro and named the dimerization initiation site (E. Skripkin, J.-C. Paillart, R. Marquet, B. Ehresmann, and C. Ehresmann, Proc. Natl. Acad. Sci. USA 91: 4945-4949, 1994). To investigate the possible role of this 5' stem-loop in HIV-1 virion formation and infectivity, four mutant viruses were generated and analyzed in vivo. Results show that deletion of the stem-loop structure reduces infectivity by a factor of 10(3) whereas loop substitutions cause a decrease of 10- to 100-fold. The level of genomic RNA packaging was found to be decreased fivefold in mutants virions containing the stem-loop deletion and only twofold in the loop-substituted virions. Surprisingly, the second DNA strand transfer during reverse transcription was found to be severely impaired upon stem-loop deletion. Taken together, these results indicate that the stem-loop structure called the dimerization initiation site is a cis element acting on both genomic RNA packaging and synthesis of proviral DNA.

192 citations

Journal ArticleDOI
01 Nov 1977-Cell
TL;DR: Six restriction fragments of Ad2 DNA which contain sites for RNA initiation have been identified by their ability to hybridize nascent labeled RNA less than 1 kb in length.

192 citations

Journal ArticleDOI
26 Jan 1996-Cell
TL;DR: Kinetic analysis shows that RNA polymerase recycling on preassembled tDNA.TFIIIB complexes is much faster than the initial transcription cycle, and template competition assays show that RNA pol III is committed to reinitiate on the same gene.

192 citations

Journal ArticleDOI
TL;DR: A combined strategy involving bacterial expression of an nsp12 fusion protein and its in vivo cleavage to generate and purify stable SARS-CoV nsp 12 with a natural N-terminus and C-terminal hexahistidine tag and possesses robust in vitro RdRp activity, as well as a significant DNA-dependent activity that may facilitate future inhibitor studies.
Abstract: An RNA-dependent RNA polymerase (RdRp) is the central catalytic subunit of the RNA-synthesizing machinery of all positive-strand RNA viruses. Usually, RdRp domains are readily identifiable by comparative sequence analysis, but biochemical confirmation and characterization can be hampered by intrinsic protein properties and technical complications. It is presumed that replication and transcription of the approximately 30-kb severe acute respiratory syndrome (SARS) coronavirus (SARS-CoV) RNA genome are catalyzed by an RdRp domain in the C-terminal part of nonstructural protein 12 (nsp12), one of 16 replicase subunits. However, thus far full-length nsp12 has proven refractory to expression in bacterial systems, which has hindered both the biochemical characterization of coronavirus RNA synthesis and RdRp-targeted antiviral drug design. Here, we describe a combined strategy involving bacterial expression of an nsp12 fusion protein and its in vivo cleavage to generate and purify stable SARS-CoV nsp12 (106 kDa) with a natural N-terminus and C-terminal hexahistidine tag. This recombinant protein possesses robust in vitro RdRp activity, as well as a significant DNA-dependent activity that may facilitate future inhibitor studies. The SARS-CoV nsp12 is primer dependent on both homo- and heteropolymeric templates, supporting the likeliness of a close enzymatic collaboration with the intriguing RNA primase activity that was recently proposed for coronavirus nsp8.

192 citations

Journal ArticleDOI
TL;DR: The genomic RNA of the avian influenza A virus, fowl plague, was fractionated into eight species by electrophoresis in polyacrylamide-agarose gels containing 6 M urea and it was demonstrated that each species has a distinct nucleotide sequence.
Abstract: The genomic RNA of the avian influenza A virus, fowl plague, was fractionated into eight species by electrophoresis in polyacrylamide-agarose gels containing 6 M urea. The separated 32P-labeled RNA species were characterized by digestion with RNase T1 and fractionation of the resulting oligonucleotides by two-dimensional gel electrophoresis; this demonstrated that each species has a distinct nucleotide sequence. A tentative correlation of each genome RNA species with the virus protein that it encodes was made.

191 citations


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Performance
Metrics
No. of papers in the topic in previous years
YearPapers
202358
2022201
2021222
2020200
2019116
2018118