Insights into RNA unwinding and ATP hydrolysis by the flavivirus NS3 protein

Authors

  • Dahai Luo,

    1. Structural & Computational Biology Division, School of Biological Sciences, Nanyang Technological University, Singapore
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    • These authors contributed equally to this work
  • Ting Xu,

    1. Dengue Unit, Novartis Institute for Tropical Diseases, Singapore
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    • These authors contributed equally to this work
  • Randall P Watson,

    1. Novartis Institutes for BioMedical Research, Discovery Technologies, Basel, Switzerland
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    • These authors contributed equally to this work
  • Daniella Scherer-Becker,

    1. Novartis Institutes for BioMedical Research, Discovery Technologies, Basel, Switzerland
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  • Aruna Sampath,

    1. Dengue Unit, Novartis Institute for Tropical Diseases, Singapore
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  • Wolfgang Jahnke,

    1. Novartis Institutes for BioMedical Research, Discovery Technologies, Basel, Switzerland
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  • Sui Sum Yeong,

    1. Structural & Computational Biology Division, School of Biological Sciences, Nanyang Technological University, Singapore
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  • Chern Hoe Wang,

    1. Structural & Computational Biology Division, School of Biological Sciences, Nanyang Technological University, Singapore
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  • Siew Pheng Lim,

    1. Dengue Unit, Novartis Institute for Tropical Diseases, Singapore
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  • Alex Strongin,

    1. Burnham Institute, La Jolla, CA, USA
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  • Subhash G Vasudevan,

    Corresponding author
    1. Program for Emerging Infectious Diseases, Duke-NUS Graduate Medical School Singapore, Singapore
    • Corresponding authors: Program for Emerging Infectious diseases, Duke-NUS Graduate Medical School Singapore, Singapore. Tel.: +65 6516 6718; Fax: +65 6534 8632; E-mail: gmsvsg@nus.edu.sgor School of Biological Sciences, Nanyang Technological University, 60 Nanyang Drive, Singapore 637551, Singapore. Tel.: +65 6316 2859; Fax: +65 6791 3856; E-mail: Julien@ntu.edu.sg

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  • Julien Lescar

    Corresponding author
    1. Structural & Computational Biology Division, School of Biological Sciences, Nanyang Technological University, Singapore
    2. Dengue Unit, Novartis Institute for Tropical Diseases, Singapore
    • Corresponding authors: Program for Emerging Infectious diseases, Duke-NUS Graduate Medical School Singapore, Singapore. Tel.: +65 6516 6718; Fax: +65 6534 8632; E-mail: gmsvsg@nus.edu.sgor School of Biological Sciences, Nanyang Technological University, 60 Nanyang Drive, Singapore 637551, Singapore. Tel.: +65 6316 2859; Fax: +65 6791 3856; E-mail: Julien@ntu.edu.sg

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Abstract

Together with the NS5 polymerase, the NS3 helicase has a pivotal function in flavivirus RNA replication and constitutes an important drug target. We captured the dengue virus NS3 helicase at several stages along the catalytic pathway including bound to single-stranded (ss) RNA, to an ATP analogue, to a transition-state analogue and to ATP hydrolysis products. RNA recognition appears largely sequence independent in a way remarkably similar to eukaryotic DEAD box proteins Vasa and eIF4AIII. On ssRNA binding, the NS3 enzyme switches to a catalytic-competent state imparted by an inward movement of the P-loop, interdomain closure and a change in the divalent metal coordination shell, providing a structural basis for RNA-stimulated ATP hydrolysis. These structures demonstrate for the first time large quaternary changes in the flaviviridae helicase, identify the catalytic water molecule and point to a β-hairpin that protrudes from subdomain 2, as a critical element for dsRNA unwinding. They also suggest how NS3 could exert an effect as an RNA-anchoring device and thus participate both in flavivirus RNA replication and assembly.

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