Analysis of the nucleoside triphosphatase, RNA triphosphatase, and unwinding activities of the helicase domain of dengue virus NS3 protein

FEBS Letters ◽  
2009 ◽  
Vol 583 (4) ◽  
pp. 691-696 ◽  
Author(s):  
Chun-Chung Wang ◽  
Zhi-Shun Huang ◽  
Pei-Ling Chiang ◽  
Chien-Tsun Chen ◽  
Huey-Nan Wu
Keyword(s):  
Dengue Virus ◽  
Helicase Domain ◽  
Ns3 Protein ◽  
Rna Triphosphatase ◽  
Nucleoside Triphosphatase

scholarly journals HCV NS3 protein helicase domain assists RNA structure conversion

FEBS Letters ◽  
2010 ◽  
Vol 584 (11) ◽  
pp. 2356-2362 ◽  
Author(s):  
Zhi-Shun Huang ◽  
Chun-Chung Wang ◽  
Huey-Nan Wu
Keyword(s):  
Rna Structure ◽  
Helicase Domain ◽  
Ns3 Protein

scholarly journals The Arginine-1493 Residue in QRRGRTGR1493G Motif IV of the Hepatitis C Virus NS3 Helicase Domain Is Essential for NS3 Protein Methylation by the Protein Arginine Methyltransferase 1

2001 ◽  
Vol 75 (17) ◽  
pp. 8031-8044 ◽  
Author(s):  
Jaerang Rho ◽  
Seeyoung Choi ◽  
Young Rim Seong ◽  
Joonho Choi ◽  
Dong-Soo Im
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Rna Binding ◽  
Rna Helicase ◽  
Full Length ◽  
Arginine Residue ◽  
Protein Methylation ◽  
Helicase Domain ◽  
Rna Helicases ◽  
Ns3 Protein

ABSTRACT The NS3 protein of hepatitis C virus (HCV) contains protease and RNA helicase activities, both of which are likely to be essential for HCV propagation. An arginine residue present in the arginine-glycine (RG)-rich region of many RNA-binding proteins is posttranslationally methylated by protein arginine methyltransferases (PRMTs). Amino acid sequence analysis revealed that the NS3 protein contains seven RG motifs, including two potential RG motifs in the 1486-QRRGRTGRG-1494 motif IV of the RNA helicase domain, in which arginines are potentially methylated by PRMTs. Indeed, we found that the full-length NS3 protein is arginine methylated in vivo. The full-length NS3 protein and the NS3 RNA helicase domain were methylated by a crude human cell extract. The purified PRMT1 methylated the full-length NS3 and the RNA helicase domain, but not the NS3 protease domain. The NS3 helicase bound specifically and comigrated with PRMT1 in vitro. Mutational analyses indicate that the Arg1493 in the QRR1488GRTGR1493G region of the NS3 RNA helicase is essential for NS3 protein methylation and that Arg1488 is likely methylated. NS3 protein methylation by the PRMT1 was decreased in the presence of homoribopolymers, suggesting that the arginine-rich motif IV is involved in RNA binding. The results suggest that an arginine residue(s) in QRXGRXGR motif IV conserved in the virus-encoded RNA helicases can be posttranslationally methylated by the PRMT1.

scholarly journals Enzymatic Defects of the nsP2 Proteins of Semliki Forest Virus Temperature-Sensitive Mutants

2007 ◽  
Vol 81 (6) ◽  
pp. 2849-2860 ◽  
Author(s):  
Giuseppe Balistreri ◽  
Javier Caldentey ◽  
Leevi Kääriäinen ◽  
Tero Ahola
Keyword(s):  
Protease Activity ◽  
Semliki Forest Virus ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Helicase Domain ◽  
Permissive Temperature ◽  
Protease Domain ◽  
Rna Triphosphatase ◽  
Interdomain Interactions

ABSTRACT We have analyzed the biochemical consequences of mutations that affect viral RNA synthesis in Semliki Forest virus temperature-sensitive (ts) mutants. Of the six mutations mapping in the multifunctional replicase protein nsP2, three were located in the N-terminal helicase region and three were in the C-terminal protease domain. Wild-type and mutant nsP2s were expressed, purified, and assayed for nucleotide triphosphatase (NTPase), RNA triphosphatase (RTPase), and protease activities in vitro at 24°C and 35°C. The protease domain mutants (ts4, ts6, and ts11) had reduced protease activity at 35°C but displayed normal NTPase and RTPase. The helicase domain mutation ts1 did not have enzymatic consequences, whereas ts13a and ts9 reduced both NTPase and protease activities but in different and mutant-specific ways. The effects of these helicase domain mutants on protease function suggest interdomain interactions within nsP2. NTPase activity was not directly required for protease activity. The similarities of the NTPase and RTPase results, as well as competition experiments, suggest that these two reactions utilize the same active site. The mutations were also studied in recombinant viruses first cultivated at the permissive temperature and then shifted up to the restrictive temperature. Processing of the nonstructural polyprotein was generally retarded in cells infected with viruses carrying the ts4, ts6, ts11, and ts13a mutations, and a specific defect appeared in ts9. All mutations except ts13a were associated with a large reduction in the production of the subgenomic 26S mRNA, indicating that both protease and helicase domains influence the recognition of the subgenomic promoter during virus replication.

Antigenicity of a recombinant NS3 protein representative of ATPase/helicase domain from hepatitis C virus

2003 ◽  
Vol 36 (1) ◽  
pp. 41-49 ◽  
Author(s):  
N Pentón ◽  
A Musacchio ◽  
J.M Rivera ◽  
J Roca ◽  
M Ponce ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Helicase Domain ◽  
Ns3 Protein

The study on the immune response induced by expressing recombinant plasmid of dengue virus type 2 NS3 protein

2000 ◽  
Vol 5 (2) ◽  
pp. 245-248
Author(s):  
Li Xiang-qun ◽  
Mao Lin ◽  
Yan Zhan-qiu ◽  
Jiang Li-feng ◽  
Yan Hui-jun ◽  
...  
Keyword(s):  
Immune Response ◽  
Dengue Virus ◽  
Virus Type ◽  
Recombinant Plasmid ◽  
Ns3 Protein ◽  
Dengue Virus Type 2

scholarly journals Helicase Domain of West Nile Virus NS3 Protein Plays a Role in Inhibition of Type I Interferon Signalling

Viruses ◽  
2017 ◽  
Vol 9 (11) ◽  
pp. 326 ◽  
Author(s):  
Yin Setoh ◽  
Parthiban Periasamy ◽  
Nias Peng ◽  
Alberto Amarilla ◽  
Andrii Slonchak ◽  
...  
Keyword(s):  
West Nile Virus ◽  
Type I Interferon ◽  
Helicase Domain ◽  
Type I ◽  
West Nile ◽  
Ns3 Protein ◽  
Interferon Signalling

scholarly journals RNA 5′-Triphosphatase Activity of the Hepatitis E Virus Helicase Domain

2010 ◽  
Vol 84 (18) ◽  
pp. 9637-9641 ◽  
Author(s):  
Yogesh A. Karpe ◽  
Kavita S. Lole
Keyword(s):  
Hepatitis E Virus ◽  
Hepatitis E ◽  
Open Reading Frame ◽  
Helicase Domain ◽  
Reading Frame ◽  
Essential Step ◽  
Positive Sense ◽  
Rna Triphosphatase ◽  
Rna Duplexes ◽  
Open Reading Frame 1

ABSTRACT Hepatitis E virus (HEV) has a positive-sense RNA genome with a 5′-m7G cap. HEV open reading frame 1 (ORF1) encodes a polyprotein with multiple enzyme domains required for replication. HEV helicase is a nucleoside triphosphatase (NTPase) with the ability to unwind RNA duplexes in the 5′-to-3′ direction. When incubated with 5′-[γ-32P]RNA and 5′-[α-32P]RNA, HEV helicase released 32P only from 5′-[γ-32P]RNA, showing specificity for the γ-β-triphosphate bond. Removal of γ-phosphate from the 5′ end of the primary transcripts (pppRNA to ppRNA) by RNA triphosphatase is an essential step during cap formation. It is suggested that HEV employs the helicase to mediate the first step of 5′ cap synthesis.

scholarly journals Chemical Mutagenesis of Dengue Virus Type 4 Yields Mutant Viruses Which Are Temperature Sensitive in Vero Cells or Human Liver Cells and Attenuated in Mice

2001 ◽  
Vol 75 (20) ◽  
pp. 9731-9740 ◽  
Author(s):  
Joseph E. Blaney ◽  
Daniel H. Johnson ◽  
Cai-Yen Firestone ◽  
Christopher T. Hanson ◽  
Brian R. Murphy ◽  
...  
Keyword(s):  
Dengue Virus ◽  
Virus Type ◽  
Human Liver ◽  
Vero Cells ◽  
Chemical Mutagenesis ◽  
Temperature Sensitive ◽  
Nucleotide Position ◽  
Nucleotide Substitutions ◽  
Ns3 Protein ◽  
Old Mice

ABSTRACT A recombinant live attenuated dengue virus type 4 (DEN4) vaccine candidate, 2AΔ30, was found previously to be generally well tolerated in humans, but a rash and an elevation of liver enzymes in the serum occurred in some vaccinees. 2AΔ30, a non-temperature-sensitive (non-ts) virus, contains a 30-nucleotide deletion (Δ30) in the 3′ untranslated region (UTR) of the viral genome. In the present study, chemical mutagenesis of DEN4 was utilized to generate attenuating mutations which may be useful in further attenuation of the 2AΔ30 candidate vaccine. Wild-type DEN4 2A virus was grown in Vero cells in the presence of 5-fluorouracil, and a panel of 1,248 clones were isolated. Twenty ts mutant viruses were identified that were ts in both simian Vero and human liver HuH-7 cells (n = 13) or only in HuH-7 cells (n = 7). Each of the 20 ts mutant viruses possessed an attenuation phenotype, as indicated by restricted replication in the brains of 7-day-old mice. The complete nucleotide sequence of the 20 ts mutant viruses identified nucleotide substitutions in structural and nonstructural genes as well as in the 5′ and 3′ UTRs, with more than one change occurring, in general, per mutant virus. A ts mutation in the NS3 protein (nucleotide position 4995) was introduced into a recombinant DEN4 virus possessing the Δ30 deletion, thereby creating rDEN4Δ30-4995, a recombinant virus which is ts and more attenuated than rDEN4Δ30 virus in the brains of mice. We are assembling a menu of attenuating mutations that should be useful in generating satisfactorily attenuated recombinant dengue vaccine viruses and in increasing our understanding of the pathogenesis of dengue virus.

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