Expression and characterization of the hepatitis G virus helicase

Laxton, Carl; McMillan, David; Ackrill, V. Sullivan A. M.

doi:10.1046/j.1365-2893.1998.00084.x

Cited by 13 publications

(5 citation statements)

References 16 publications

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“…On the basis of sequence motif analysis, the HGV NS3 protein is predictive of a multifunctional protein containing serine protease, ATPase, and RNA helicase activities (2,14). Recently, it was found that the HGV NS3 protein has the ATPase and DNA helicase activities (12). In this study we confirm that the HGV NS3 protein has an RNA-stimulated ATPase activity and an RNA helicase activity, and we characterize the biochemical properties of the protein in detail.…”

Section: Discussionsupporting

confidence: 68%

RNA-Stimulated ATPase and RNA Helicase Activities and RNA Binding Domain of Hepatitis G Virus Nonstructural Protein 3

Gwack

Yoo

Song

et al. 1999

J Virol

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Hepatitis G virus (HGV) nonstructural protein 3 (NS3) contains amino acid sequence motifs typical of ATPase and RNA helicase proteins. In order to examine the RNA helicase activity of the HGV NS3 protein, the NS3 region (amino acids 904 to 1580) was fused with maltose-binding protein (MBP), and the fusion protein was expressed inEscherichia coli and purified with amylose resin and anion-exchange chromatography. The purified MBP-HGV/NS3 protein possessed RNA-stimulated ATPase and RNA helicase activities. Characterization of the ATPase and RNA helicase activities of MBP-HGV/NS3 showed that the optimal reaction conditions were similar to those of other Flaviviridae viral NS3 proteins. However, the kinetic analysis of NTPase activity showed that the MBP-HGV/NS3 protein had several unique properties compared to the otherFlaviviridae NS3 proteins. The HGV NS3 helicase unwinds RNA-RNA duplexes in a 3′-to-5′ direction and can unwind RNA-DNA heteroduplexes and DNA-DNA duplexes as well. In a gel retardation assay, the MBP-HGV/NS3 helicase bound to RNA, RNA/DNA, and DNA duplexes with 5′ and 3′ overhangs but not to blunt-ended RNA duplexes. We also found that the conserved motif VI was important for RNA binding. Further deletion mapping showed that the RNA binding domain was located between residues 1383 and 1395, QRRGRTGRGRSGR. Our data showed that the MBP-HCV/NS3 protein also contains the RNA binding domain in the similar domain.

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Section: Discussionsupporting

confidence: 68%

RNA-Stimulated ATPase and RNA Helicase Activities and RNA Binding Domain of Hepatitis G Virus Nonstructural Protein 3

Gwack

Yoo

Song

et al. 1999

J Virol

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“…To date, however, the RNA helicase activity of flavivirus was demonstrated only in the NS3 protein of dengue virus type 2 [4]. The RNA‐stimulated NTPase and RNA helicase activities were also identified in the NS3 proteins of viruses which are genetically related to flavivirus such as hepatitis C virus (HCV), hepatitis G virus (HGV) and bovine viral diarrhea virus, a pestivirus (BVDV) [10–19]. The roles of such enzymatic activities of the flavivirus NS3 protein in viral replication are poorly understood.…”

Section: Introductionmentioning

confidence: 99%

Identification and characterization of the RNA helicase activity of Japanese encephalitis virus NS3 protein

Utama¹,

Shimizu²,

Morikawa³

et al. 1999

FEBS Letters

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The NS3 protein of Japanese encephalitis virus (JEV) contains motifs typical of RNA helicase/NTPase but no RNA helicase activity has been reported for this protein. To identify and characterize the RNA helicase activity of JEV NS3, a truncated form of the protein with a His-tag was expressed in Escherichia coli and purified. The purified JEV NS3 protein showed an RNA helicase activity, which was dependent on divalent cations and ATP. An Asp-285-to-Ala substitution in motif II of the JEV NS3 protein abolished the ATPase and RNA helicase activities. These results indicate that the C-terminal 457 residues are sufficient to exhibit the RNA helicase activity of JEV NS3.z 2000 Federation of European Biochemical Societies.

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“…Inhibitors of the helicase of Herpes simplex virus have been identified (208). The HCV (69,189) and HGV (134), helicases have been cloned, expressed, and characterized. The helicase of the Flaviviridae belongs to the DEAD (Asp-Glu-Ala-Asp) box family of RNA helicases (115).…”

Section: Inhibition Of Helicasementioning

confidence: 99%