Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.7.48 (transcriptase)
 9,479 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The RNA-dependent RNA polymerase L protein of vesicular stomatitis virus (VSV) elicits GTPase and RNA:GDP polyribonucleotidyltransferase (PRNTase) activities to produce a 5'-cap core structure, guanosine(5')triphospho(5')adenosine (GpppA), on viral mRNAs. Here, we report that the L protein produces an unusual cap structure, guanosine(5')tetraphospho(5')adenosine (GppppA), that is formed by the transfer of the 5'-monophosphorylated viral mRNA start sequence to GTP by the PRNTase activity before the removal of the gamma-phosphate from GTP by GTPase. Interestingly, GppppA-capped and polyadenylated full-length mRNAs were also found to be synthesized by an in vitro transcription system with the native VSV RNP.
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PMID:Formation of guanosine(5')tetraphospho(5')adenosine cap structure by an unconventional mRNA capping enzyme of vesicular stomatitis virus. 1849 67

The influenza virus polymerase is formed by the PB1, PB2 and PA subunits and is required for virus transcription and replication in the nucleus of infected cells. As PB2 is a relevant host-range determinant we expressed a TAP-tagged PB2 in human cells and isolated intracellular complexes. Alpha-importin was identified as a PB2-associated factor by proteomic analyses. To study the relevance of this interaction for virus replication we mutated the PB2 NLS and analysed the phenotype of mutant subunits, polymerase complexes and RNPs. While mutant PB2 proteins showed reduced nuclear accumulation, they formed polymerase complexes normally when co expressed with PB1 and PA. However, mutant RNPs generated with a viral CAT replicon showed up to hundred-fold reduced CAT accumulation. Rescue of nuclear localisation of mutant PB2 by insertion of an additional SV40 TAg-derived NLS did not revert the mutant phenotype of RNPs. Furthermore, determination of recombinant RNP accumulation in vivo indicated that PB2 NLS mutations drastically reduced virus RNA replication. These results indicate that, above and beyond its role in nuclear accumulation, PB2 interaction with alpha-importins is required for virus RNA replication. To ascertain whether PB2-alpha-importin binding could contribute to the adaptation of H5N1 avian viruses to man, their association in vivo was determined. Human alpha importin isoforms associated efficiently to PB2 protein of an H3N2 human virus but bound to diminished and variable extents to PB2 from H5N1 avian or human strains, suggesting that the function of alpha importin during RNA replication is important for the adaptation of avian viruses to the human host.
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PMID:The host-dependent interaction of alpha-importins with influenza PB2 polymerase subunit is required for virus RNA replication. 1906 26

Bunyaviruses are a large family of segmented RNA viruses which, like influenza virus, use a cap-snatching mechanism for transcription whereby short capped primers derived by endonucleolytic cleavage of host mRNAs are used by the viral RNA-dependent RNA polymerase (L-protein) to transcribe viral mRNAs. It was recently shown that the cap-snatching endonuclease of influenza virus resides in a discrete N-terminal domain of the PA polymerase subunit. Here we structurally and functionally characterize a similar endonuclease in La Crosse orthobunyavirus (LACV) L-protein. We expressed N-terminal fragments of the LACV L-protein and found that residues 1-180 have metal binding and divalent cation dependent nuclease activity analogous to that of influenza virus endonuclease. The 2.2 A resolution X-ray crystal structure of the domain confirms that LACV and influenza endonucleases have similar overall folds and identical two metal binding active sites. The in vitro activity of the LACV endonuclease could be abolished by point mutations in the active site or by binding 2,4-dioxo-4-phenylbutanoic acid (DPBA), a known influenza virus endonuclease inhibitor. A crystal structure with bound DPBA shows the inhibitor chelating two active site manganese ions. The essential role of this endonuclease in cap-dependent transcription was demonstrated by the loss of transcriptional activity in a RNP reconstitution system in cells upon making the same point mutations in the context of the full-length LACV L-protein. Using structure based sequence alignments we show that a similar endonuclease almost certainly exists at the N-terminus of L-proteins or PA polymerase subunits of essentially all known negative strand and cap-snatching segmented RNA viruses including arenaviruses (2 segments), bunyaviruses (3 segments), tenuiviruses (4-6 segments), and orthomyxoviruses (6-8 segments). This correspondence, together with the well-known mapping of the conserved polymerase motifs to the central regions of the L-protein and influenza PB1 subunit, suggests that L-proteins might be architecturally, and functionally equivalent to a concatemer of the three orthomyxovirus polymerase subunits in the order PA-PB1-PB2. Furthermore, our structure of a known influenza endonuclease inhibitor bound to LACV endonuclease suggests that compounds targeting a potentially broad spectrum of segmented RNA viruses, several of which are serious or emerging human, animal and plant pathogens, could be developed using structure-based optimisation.
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PMID:Bunyaviridae RNA polymerases (L-protein) have an N-terminal, influenza-like endonuclease domain, essential for viral cap-dependent transcription. 2086 19

All representatives of rhabdoviruses contain a nucleocapside phosphoprotein - P-protein which is an essential subunit of the viral RNA-dependent RNA polymerase complex. As a result of studying the effect of nucleocapside protein P(NS) on replicase activity of mRNP isolated from plants infected by potato curly dwarf virus in the system in vitro, it was established that nucleocapside P-protein stimulates considerably the replicase activity of membrane-bound polysomal m-RNP P-protein being available in concentration of 15 microg/ml in the replication system in vitro of membrane-bound polysomal mRNP, the replicase activity increased 11.7 times. This property of nucleocapside P-protein at the same concentration was displayed to a less extent with the presence of free polysomal mRNP, in the system in vitro. Thus the replicase activity mRNP-complexes in the replication system in vitro depends on the presence of nucleocapside viral P-protein in the system. Its concentration being increased or decreased, one can observe the change of the replicase activity.
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PMID:[Effect of structural P-protein of potato curly dwarf virus on replicase activity of mRNP-complexes of infected plants]. 2230 54


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