scholarly journals Analysis of tombusvirus revertants to identify essential amino acid residues within RNA-dependent RNA polymerase motifs

2005 ◽  
Vol 86 (3) ◽  
pp. 823-826 ◽  
Author(s):  
K. Boonrod ◽  
S. Chotewutmontri ◽  
D. Galetzka ◽  
G. Krczal
Keyword(s):  
Rna Polymerase ◽  
Nicotiana Benthamiana ◽  
Essential Amino Acid ◽  
Selection Pressure ◽  
Rna Binding ◽  
Amino Acid Residues ◽  
Rna Dependent Rna Polymerase ◽  
Viral Rdrp ◽  
In Trans ◽  
Residue Position

The RNA-dependent RNA polymerase (RdRp) of Tomato bushy stunt virus (TBSV) contains an arginine- and proline-rich (RPR) motif. This motif functions as an RNA-binding domain and is essential for tombusvirus replication. A mutant carrying three arginine substitutions in this motif rendered the virus unable to replicate in Nicotiana benthamiana plants and protoplasts. When the replicase function was provided in trans, by expressing the TBSV RdRp in N. benthamiana plants, an infectious variant could be isolated. Sequence analysis showed that only the substituted glycine residue (position 216) had reverted to arginine; all other substitutions remained unchanged. This finding suggested that strong selection pressure is active to maintain necessary sequences of the viral RdRp and that the analysis of revertants may help to identify essential viral functions.

scholarly journals Conformational changes in human Norovirus polymerase

2014 ◽  
Vol 70 (a1) ◽  
pp. C1595-C1595
Author(s):  
Kenneth Ng ◽  
Dmitry Zamyatkin ◽  
Hayeong Rho ◽  
Elesha Hoffarth ◽  
Gabriela Jurca ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Conformational Changes ◽  
Rna Binding ◽  
Divalent Metal ◽  
Rna Dependent Rna Polymerase ◽  
Human Norovirus ◽  
Conformational States ◽  
Crystal Forms ◽  
Divalent Metal Cations ◽  
Positive Strand Rna

Human Noroviruses (NV) belong in the Caliciviridae family and are a major cause of gastroenteritis outbreaks throughout the world. Crystal structures of the RNA-dependent RNA polymerase from the human Norovirus have been determined in over ten different crystal forms in the presence and absence of divalent metal cations, nucleoside triphosphates, inhibitors and primer-template duplex RNA. These structures show how the polymerase enzyme can adopt a range of conformations in which the thumb, fingers and palm domains change orientations depending on the step of the enzymatic cycle trapped in different crystal forms. We discuss how the evidence from crystallographic and biochemical experiments combine to better understand how viral RNA polymerase enzymes from human Norovirus and related positive-strand RNA viruses can adopt a range of conformational states to facilitate RNA binding, NTP binding, catalysis, RNA translocation and pyrophosphate release. The detailed structural and mechanistic understanding of these conformational changes is important for providing a sound basis for understanding viral replication in general, as well as for the design of novel inhibitors capable of trapping the enzyme in specific conformational states.

scholarly journals Sequences at the 3′ Untranslated Region of Bamboo Mosaic Potexvirus RNA Interact with the Viral RNA-Dependent RNA Polymerase

2001 ◽  
Vol 75 (6) ◽  
pp. 2818-2824 ◽  
Author(s):  
Cheng-Yen Huang ◽  
Yih-Leh Huang ◽  
Menghsiao Meng ◽  
Yau-Heiu Hsu ◽  
Ching-Hsiu Tsai
Keyword(s):  
Rna Polymerase ◽  
Untranslated Region ◽  
Rna Binding ◽  
Bovine Liver ◽  
Viral Rna ◽  
Mobility Shift ◽  
Rna Dependent Rna Polymerase ◽  
Stem Loop ◽  
Competition Analysis ◽  
Reading Frame

ABSTRACT The 3′ untranslated region (UTR) of bamboo mosaic potexvirus (BaMV) genomic RNA was found to fold into a series of stem-loop structures including a pseudoknot structure. These structures were demonstrated to be important for viral RNA replication and were believed to be recognized by the replicase (C.-P. Cheng and C.-H. Tsai, J. Mol. Biol. 288:555–565, 1999). Electrophoretic mobility shift and competition assays have now been used to demonstrate that theEscherichia coli-expressed RNA-dependent RNA polymerase domain (Δ893) derived from BaMV open reading frame 1 could specifically bind to the 3′ UTR of BaMV RNA. No competition was observed when bovine liver tRNAs or poly(I)(C) double-stranded homopolymers were used as competitors, and the cucumber mosaic virus 3′ UTR was a less efficient competitor. Competition analysis with different regions of the BaMV 3′ UTR showed that Δ893 binds to at least two independent RNA binding sites, stem-loop D and the poly(A) tail. Footprinting analysis revealed that Δ893 could protect the sequences at loop D containing the potexviral conserved hexamer motif and part of the stem of domain D from chemical cleavage.

scholarly journals Identification of Residues Critical for the Interferon Antagonist Function of Langat Virus NS5 Reveals a Role for the RNA-Dependent RNA Polymerase Domain

2007 ◽  
Vol 81 (13) ◽  
pp. 6936-6946 ◽  
Author(s):  
Gregory S. Park ◽  
Keely L. Morris ◽  
Roselyn G. Hallett ◽  
Marshall E. Bloom ◽  
Sonja M. Best
Keyword(s):  
Amino Acids ◽  
Rna Polymerase ◽  
Nonstructural Protein ◽  
Encephalitis Virus ◽  
Janus Kinase ◽  
Amino Acid Residues ◽  
Rna Dependent Rna Polymerase ◽  
Stat Signaling ◽  
Langat Virus ◽  
Tick Borne Encephalitis Virus

ABSTRACT All pathogenic flaviviruses examined thus far inhibit host interferon (IFN) responses by suppressing the Janus kinase-signal transducer and activator of transcription (JAK-STAT) pathway. Both Langat virus (LGTV; a member of the tick-borne encephalitis virus serogroup) and Japanese encephalitis virus use the nonstructural protein NS5 to suppress JAK-STAT signaling. However, NS5 is also critical to virus replication, contributing methyltransferase and RNA-dependent RNA polymerase (RdRP) activities. The specific amino acid residues of NS5 involved in IFN antagonism are not known. Here, we demonstrate that the LGTV NS5 JAK-STAT inhibitory domain is contained between amino acids 355 and 735 (of 903), a range which lies within the RdRP domain. Furthermore, we identified two noncontiguous stretches of specific amino acids within the RdRP, 374 to 380 and 624 to 647, as critical for inhibition of JAK-STAT signaling. Despite considerable separation on the linear NS5 sequence, these residues localized adjacent to each other when modeled on the West Nile virus RdRP crystal structure. Due to the general conservation of RdRP structures, these results suggest that the specific residues identified act cooperatively to form a unique functional site on the RdRP responsible for JAK-STAT inhibition. This insight into the mechanism underlying flavivirus IFN evasion strategies will facilitate the design of antiviral therapeutics that potentiate the action of IFN during infection.

scholarly journals RNA-Dependent RNA Polymerase 1 from Nicotiana tabacum Suppresses RNA Silencing and Enhances Viral Infection in Nicotiana benthamiana

2010 ◽  
Vol 22 (4) ◽  
pp. 1358-1372 ◽  
Author(s):  
Xiao-Bao Ying ◽  
Li Dong ◽  
Hui Zhu ◽  
Cheng-Guo Duan ◽  
Quan-Sheng Du ◽  
...  
Keyword(s):  
Nicotiana Tabacum ◽  
Rna Polymerase ◽  
Viral Infection ◽  
Nicotiana Benthamiana ◽  
Rna Silencing ◽  
Rna Dependent Rna Polymerase ◽  
Rna Polymerase 1

scholarly journals Functional Analysis of RNA Binding by the Hepatitis C Virus RNA-dependent RNA Polymerase

2005 ◽  
Vol 280 (45) ◽  
pp. 38011-38019 ◽  
Author(s):  
Young-Chan Kim ◽  
William K. Russell ◽  
C. T. Ranjith-Kumar ◽  
Michael Thomson ◽  
David H. Russell ◽  
...  
Keyword(s):  
Functional Analysis ◽  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Rna Polymerase ◽  
Rna Binding ◽  
Hepatitis C Virus Rna ◽  
Rna Dependent Rna Polymerase ◽  
Virus Rna

scholarly journals Inhibition of RNA binding to hepatitis C virus RNA-dependent RNA polymerase: a new mechanism for antiviral intervention

2014 ◽  
Vol 42 (14) ◽  
pp. 9399-9409 ◽  
Author(s):  
Abdelhakim Ahmed-Belkacem ◽  
Jean-François Guichou ◽  
Rozenn Brillet ◽  
Nazim Ahnou ◽  
Eva Hernandez ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Rna Polymerase ◽  
Rna Binding ◽  
Hepatitis C Virus Rna ◽  
Rna Dependent Rna Polymerase ◽  
Virus Rna

scholarly journals Potential Phytochemical Inhibitors of the Coronavirus RNA Dependent RNA Polymerase: A Molecular Docking Study

2020 ◽  
Author(s):  
Ardra. P ◽  
Prachi Singh ◽  
Hariprasad VR ◽  
Babu UV ◽  
Mohamed Rafiq ◽  
...  
Keyword(s):  
Molecular Docking ◽  
Rna Polymerase ◽  
Metal Ion ◽  
Docking Study ◽  
Nucleoside Triphosphate ◽  
Root Extract ◽  
Amino Acid Residues ◽  
Molecular Docking Study ◽  
Rna Dependent Rna Polymerase ◽  
Phytochemical Compounds

Abstract The COVID-19 disease that originated in China by the end of 2019 has now become a pandemic and has affected 216 countries as on 08 June 2020. RNA dependent RNA polymerase (RdRp), the core enzyme in the multiprotein replicase-transcriptase complex of coronaviruses, serves as a classical target for inhibiting the coronavirus infectivity. In this study we performed molecular docking of sixty-nine different phytochemical compounds from various herbs with RdRp of both SARS-CoV-2 and its predecessor SARS-CoV. Our results show that various phytochemical constituents from Withania somnifera root extract, Hyssopus officinalis and Camellia sinensis leaf extract have high binding affinity towards RdRps and are comparable to the small molecule drug remdesivir. Their binding interactions reveal that they bind to the amino acid residues involved in nucleoside triphosphate (NTP) entry and recognition site and metal ion cofactor chelating site of both SARS-CoV-2 and SARS-CoV. Hence they are different from the classical nucleotide analog inhibitors of RdRp. This study paves a quick platform for development of targeted therapy using phytochemicals for COVID-19 and other potential SARS coronavirus related outbreaks in future.

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