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Journal Articles Journal of Virology Year : 2016

Domain organization of vaccinia virus helicase-primase D5.


Poxviridae are viruses with a large linear double-stranded DNA genome coding for up to 250 open reading frames and a fully cytoplasmic replication. The double-stranded DNA genome is covalently circularized at both ends and is replicated fully in the cytoplasm. Similar structures of covalently linked extremities of the linear DNA genome are found in the African swine fever virus (asfarvirus) and in phycodnaviridae. We are studying the machinery which replicates this peculiar genome structure. Working on vaccinia virus we give first insights into the overall structure and function of the essential poxvirus virus helicase-primase D5 and show that the active helicase domain of D5 builds a hexameric ring structure. This hexamer has ATPase and, more generally, nucleoside triphosphatase activity indistinguishable from full length D5 independent of the nature of the base. In addition, hexameric helicase domains bind tightly to single- and double-stranded DNA. Still, the monomeric D5 helicase construct truncated within the D5N domain leads to a well-defined structure, but without ATPase or DNA binding activity. This shows that the full D5N domain has to be present for hexamerisation. This allowed us to assign a function the D5N domain which is not only present in D5, but also in other viruses of the Nucleo-Cytoplasmic Large DNA Virus (NCLDV) clade. The primase domain and the helicase domain were structurally analyzed via a combination of SAXS and, when appropriate, electron microscopy, leading to consistent low-resolution models of the different proteins. Since the beginning of the 1980s, research on the vaccinia virus replication mechanism has basically stalled due to the absence of structural information. As a result this important class of pathogens is less well understood than most other viruses. This lack of information concerns in general viruses of NCLDV clade, which use a SF3 superfamily helicase for replication as poxviruses do. Here we show for the first time information about the domain structure and DNA-binding activity of D5, the poxvirus helicase-primase. This result refines not only the current model of the poxvirus replication fork but, on the long run, will lead to a structural base for anti-viral drug design.
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Dates and versions

hal-01287707 , version 1 (14-03-2016)


  • HAL Id : hal-01287707 , version 1
  • PUBMED : 26912611


Stephanie Hutin, Wai-Li Ling, Adam Round, Grégory Effantin, Stefan Reich, et al.. Domain organization of vaccinia virus helicase-primase D5.. Journal of Virology, 2016. ⟨hal-01287707⟩
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