The flavivirus polymerase as a target for drug discovery

Malet, Hélène; Massé, Nicolas; Selisko, Barbara; Romette, Jean‐Louis; Alvarez, Karine; Guillemot, Jean Claude; Tolou, H.; Yap, Thai Leong; Vasudevan, Subhash G.; Lescar, Julien; Canard, Bruno

doi:10.1016/j.antiviral.2008.06.007

Cited by 176 publications

(198 citation statements)

References 83 publications

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“…Significant nucleoside-based drug discovery and development have been undertaken and are ongoing to combat viral infections caused by Flaviviridae members, such as HCV and dengue virus (13)(14)(15)(16)(17)(18)(19). Sofosbuvir, an FDA-approved ribonucleotide analog targeting HCV NS5b polymerase, has been shown to be a very effective and safe drug for the treatment of HCV infection, and these findings provided a proof of concept for the use of ribonucleotide analogs as effective antiviral drugs against Flaviviridae (20,21).…”

mentioning

confidence: 99%

Analysis of Ribonucleotide 5′-Triphosphate Analogs as Potential Inhibitors of Zika Virus RNA-Dependent RNA Polymerase by Using Nonradioactive Polymerase Assays

Bluemling

Collop

et al. 2017

Antimicrob Agents Chemother

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Zika virus (ZIKV) is an emerging human pathogen that is spreading rapidly through the Americas and has been linked to the development of microcephaly and to a dramatically increased number of Guillain-Barré syndrome cases. Currently, no vaccine or therapeutic options for the prevention or treatment of ZIKV infections exist. In the study described in this report, we expressed, purified, and characterized full-length nonstructural protein 5 (NS5) and the NS5 polymerase domain (NS5pol) of ZIKV RNA-dependent RNA polymerase. Using purified NS5, we developed an in vitro nonradioactive primer extension assay employing a fluorescently labeled primertemplate pair. Both purified NS5 and NS5pol can carry out in vitro RNA-dependent RNA synthesis in this assay. Our results show that Mn 2ϩ is required for enzymatic activity, while Mg 2ϩ is not. We found that ZIKV NS5 can utilize single-stranded DNA but not double-stranded DNA as a template or a primer to synthesize RNA. The assay was used to compare the efficiency of incorporation of analog 5=-triphosphates by the ZIKV polymerase and to calculate their discrimination versus that of natural ribonucleotide triphosphates (rNTPs). The 50% inhibitory concentrations for analog rNTPs were determined in an alternative nonradioactive coupled-enzyme assay. We determined that, in general, 2=-C-methyl-and 2=-C-ethynyl-substituted analog 5=-triphosphates were efficiently incorporated by the ZIKV polymerase and were also efficient chain terminators. Derivatives of these molecules may serve as potential antiviral compounds to be developed to combat ZIKV infection. This report provides the first characterization of ZIKV polymerase and demonstrates the utility of in vitro polymerase assays in the identification of potential ZIKV inhibitors.

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mentioning

confidence: 99%

Analysis of Ribonucleotide 5′-Triphosphate Analogs as Potential Inhibitors of Zika Virus RNA-Dependent RNA Polymerase by Using Nonradioactive Polymerase Assays

Bluemling

Collop

et al. 2017

Antimicrob Agents Chemother

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“…The essential role of NS5 in flavivirus replication makes it an attractive antiviral target (Malet et al, 2008). Thus, the structure of the full-length molecule is required to provide a more accurate model for functional analysis and drug design.…”

Section: Introductionmentioning

confidence: 99%

Monoclonal antibodies to the West Nile virus NS5 protein map to linear and conformational epitopes in the methyltransferase and polymerase domains

Hall

Tan

Selisko

et al. 2009

Journal of General Virology

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The West Nile virus (WNV) NS5 protein contains a methyltransferase (MTase) domain involved in RNA capping and an RNA-dependent RNA polymerase (RdRp) domain essential for virus replication. Crystal structures of individual WNV MTase and RdRp domains have been solved; however, the structure of full-length NS5 has not been determined. To gain more insight into the structure of NS5 and interactions between the MTase and RdRp domains, we generated a panel of seven monoclonal antibodies (mAbs) to the NS5 protein of WNV (Kunjin strain) and mapped their binding sites using a series of truncated NS5 proteins and synthetic peptides. Binding sites of four mAbs (5D4, 4B6, 5C11 and 6A10) were mapped to residues 354-389 in the fingers subdomain of the RdRp. This is consistent with the ability of these mAbs to inhibit RdRp activity in vitro and suggests that this region represents a potential target for RdRp inhibitors. Using a series of synthetic peptides, we also identified a linear epitope (bound by mAb 5H1) that mapped to a 13 aa stretch surrounding residues 47 and 49 in the MTase domain, a region predicted to interact with the palm subdomain of the RdRp. The failure of one mAb (7G6) to bind both N-and Cterminally truncated NS5 recombinants indicates that the antibody recognizes a conformational epitope that requires the presence of residues in both the MTase and RdRp domains. These data support a structural model of the full-length NS5 molecule that predicts a physical interaction between the MTase and the RdRp domains.

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“…6 Penelitian tersebut umumnya menitik-beratkan pada proses masuknya protein virus ke dalam sel, replikasi, perakitan, dan pematangan, sehingga diketahui tahapan penting perakitan virus yang dapat dijadikan sebagai target obat. [7][8][9][10] Beberapa protein DENV yang memiliki potensi untuk dijadikan target obat ada-lah glikoprotein envelope (E), 7 protein NS3 yang berfungsi sebagai helikase, 8 dan protein NS5 yang memiliki dua fungsi sebagai metiltransferase 9 serta RNAdependent RNA polymerase. 10 DENV adalah anggota dari keluarga Flaviviridae dan dikelompokkan dalam genus flavivirus dengan genom yang terdiri atas RNA positive-sense berukuran ~ 11kb.…”

Section: Pendahuluanunclassified

“…[7][8][9][10] Beberapa protein DENV yang memiliki potensi untuk dijadikan target obat ada-lah glikoprotein envelope (E), 7 protein NS3 yang berfungsi sebagai helikase, 8 dan protein NS5 yang memiliki dua fungsi sebagai metiltransferase 9 serta RNAdependent RNA polymerase. 10 DENV adalah anggota dari keluarga Flaviviridae dan dikelompokkan dalam genus flavivirus dengan genom yang terdiri atas RNA positive-sense berukuran ~ 11kb. RNA tersebut mengkode tiga protein struktural, yaitu protein kapsid (C), premembran/ membran (prM/M), dan E yang akan membentuk komponen virion, serta tujuh protein non-struktural, yaitu NS1, NS2A/B, NS3, NS4A/B, NS5, yang terlibat dalam replikasi RNA virus (Gambar 1).…”

Section: Pendahuluanunclassified

Struktur Proteomik Virus Dengue Dan Manfaatnya Sebagai Target Antivirus

Rachmayanti¹

2015

MKA

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AbstrakVirus dengue (DENV) telah menyebabkan sekitar 50 juta kasus infeksi demam berdarah setiap tahunnya, akan tetapi hingga saat ini belum terdapat vaksin maupun antivirus yang mampu mencegah atau mengobati penyakit tersebut. Selama pengembangan vaksin dan antivirus, diperoleh berbagai informasi tentang struktur protein DENV yang dapat dimanfaatkan sebagai target obat. Makalah membahas tentang struktur proteomik pada DENV, yaitu glikoprotein pada envelope, NS3 protease, NS3 helikase, NS5 metiltransferase, dan NS5 RNA-dependent RNA polimerase.

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The flavivirus polymerase as a target for drug discovery

Cited by 176 publications

References 83 publications

Analysis of Ribonucleotide 5′-Triphosphate Analogs as Potential Inhibitors of Zika Virus RNA-Dependent RNA Polymerase by Using Nonradioactive Polymerase Assays

Analysis of Ribonucleotide 5′-Triphosphate Analogs as Potential Inhibitors of Zika Virus RNA-Dependent RNA Polymerase by Using Nonradioactive Polymerase Assays

Monoclonal antibodies to the West Nile virus NS5 protein map to linear and conformational epitopes in the methyltransferase and polymerase domains

Struktur Proteomik Virus Dengue Dan Manfaatnya Sebagai Target Antivirus

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