Flaviviral helicase: Insights into the mechanism of action of a motor protein

Mastrangelo, Eloise; Bolognesi, Martino; Milani, Mario

doi:10.1016/j.bbrc.2011.11.060

Cited by 19 publications

(20 citation statements)

References 15 publications

(28 reference statements)

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“…Structures displaying similar closed and open conformations of the apo-NS3 enzyme from HCV have also been reported, albeit from NS3 belonging to different HCV genotypes (25,(63)(64)(65). Recent molecular dynamics simulations indicated that DENV apo-NS3h also appears to oscillate between open and closed conformations (66), again suggesting an intrinsically dynamic behavior, although the structures of the apo-form of the enzyme in different conformations are not available. SAXS analyses of HCV and flavivirus NS3h in the presence or absence of an ATP analog might provide useful comparative information.…”

Section: Discussionmentioning

confidence: 82%

X-Ray Structure of the Pestivirus NS3 Helicase and Its Conformation in Solution

Tortorici

Duquerroy

Kwok

et al. 2015

J Virol

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Pestiviruses form a genus in the Flaviviridae family of small enveloped viruses with a positive-sense single-stranded RNA genome. Viral replication in this family requires the activity of a superfamily 2 RNA helicase contained in the C-terminal domain of nonstructural protein 3 (NS3). NS3 features two conserved RecA-like domains (D1 and D2) with ATPase activity, plus a third domain (D3) that is important for unwinding nucleic acid duplexes. We report here the X-ray structure of the pestivirus NS3 helicase domain (pNS3h) at a 2.5-Å resolution. The structure deviates significantly from that of NS3 of other genera in the Flaviviridae family in D3, as it contains two important insertions that result in a narrower nucleic acid binding groove. We also show that mutations in pNS3h that rescue viruses from which the core protein is deleted map to D3, suggesting that this domain may be involved in interactions that facilitate particle assembly. Finally, structural comparisons of the enzyme in different crystalline environments, together with the findings of small-angle X-ray-scattering studies in solution, show that D2 is mobile with respect to the rest of the enzyme, oscillating between closed and open conformations. Binding of a nonhydrolyzable ATP analog locks pNS3h in a conformation that is more compact than the closest apo-form in our crystals. Together, our results provide new insight and bring up new questions about pNS3h function during pestivirus replication. IMPORTANCEAlthough pestivirus infections impose an important toll on the livestock industry worldwide, little information is available about the nonstructural proteins essential for viral replication, such as the NS3 helicase. We provide here a comparative structural and functional analysis of pNS3h with respect to its orthologs in other viruses of the same family, the flaviviruses and hepatitis C virus. Our studies reveal differences in the nucleic acid binding groove that could have implications for understanding the unwinding specificity of pNS3h, which is active only on RNA duplexes. We also show that pNS3h has a highly dynamic behavior-a characteristic probably shared with NS3 helicases from all Flaviviridae members-that could be targeted for drug design by using recent algorithms to specifically block molecular motion. Compounds that lock the enzyme in a single conformation or limit its dynamic range of conformations are indeed likely to block its helicase function. P estiviruses infect a wide range of cloven-hoofed animals, wild and domestic, causing serious disease. The most studied are the classical swine fever virus (CSFV) (1) and the bovine viral diarrhea virus (BVDV), which impose important economic losses to the livestock industry worldwide (2). They form a genus within the Flaviviridae family of single-stranded RNA viruses, which also includes medically important pathogens in the flavi-and hepacivirus genera. Recent studies also revealed that hepaci-and pegiviruses (the fourth genus in the family) cause infections in horses and other d...

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Section: Discussionmentioning

confidence: 82%

X-Ray Structure of the Pestivirus NS3 Helicase and Its Conformation in Solution

Tortorici

Duquerroy

Kwok

et al. 2015

J Virol

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“…The flaviviral helicase belongs to the helicase superfamily 2 (SF2) and is located at the C-terminal domain of NS3 (REFS 152,153). NS3 is responsible for unwinding viral RNA during replication, and its activity is driven by an intrinsic NTPase activity 152,153 . The structure of NS3 helicase has been elucidated for many flaviviruses 154 , including ZIKV 155 .…”

Section: Target Assaysmentioning

confidence: 99%

Broad-spectrum agents for flaviviral infections: dengue, Zika and beyond

Boldescu

Behnam

Vasilakis³

et al. 2017

Nat Rev Drug Discov

248

217

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Infections with flaviviruses, such as dengue, West Nile virus, and the recently re-emerging Zika virus are an increasing and probably lasting global risk. This review summarizes and comments on the opportunities for broad-spectrum agents that are active against a range of flaviviruses. Broad-spectrum activity would be particularly desirable as preparatory measure for the next flaviviral epidemic that could emerge from as-yet-unknown or neglected viruses. Potential target sites for broad-spectrum anti-flaviviral compounds include viral proteins and host mechanisms that are exploited by these viruses during entry and replication. A variety of compounds with broad-spectrum antiviral activity have already been identified by target-specific or phenotypic assays. For some other compound classes, broad-spectrum activity can be anticipated because of their mode of action and molecular target(s).

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“…The order of RNA capping has not been completely defined for flaviviruses, but the canonical Ping‐Pong mechanism for cap formation appears to be the most likely scenario (Figure ). The RNA triphosphatase is located in the NS3 helicase domain, and the RNA triphosphatase appears to overlap with the NTPase active site that powers the helicase . Newly synthesized negative and positive strand RNAs are triphosphorylated and would be appropriate substrates for the RNA triphosphatase.…”

Section: Rna Capping In Flavivirus Rna Replicationmentioning

confidence: 99%

Regulation of flavivirus RNA synthesis and capping

Saeedi

Geiss

2013

WIREs RNA

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RNA viruses, such as flaviviruses, are able to efficiently replicate and cap their RNA genomes in vertebrate and invertebrate cells. Flaviviruses use several specialized proteins to first make an uncapped negative strand copy of the viral genome that is used as a template for the synthesis of large numbers of capped genomic RNAs. Despite using relatively simple mechanisms to replicate their RNA genomes, there are significant gaps in our understanding of how flaviviruses switch between negative and positive strand RNA synthesis and how RNA capping is regulated. Recent work has begun to provide a conceptual framework for flavivirus RNA replication and capping and shown some surprising roles for genomic RNA during replication and pathogenesis.

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Flaviviral helicase: Insights into the mechanism of action of a motor protein

Cited by 19 publications

References 15 publications

X-Ray Structure of the Pestivirus NS3 Helicase and Its Conformation in Solution

X-Ray Structure of the Pestivirus NS3 Helicase and Its Conformation in Solution

Broad-spectrum agents for flaviviral infections: dengue, Zika and beyond

Regulation of flavivirus RNA synthesis and capping

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