title: Insights into RNA unwinding and ATP hydrolysis by the flavivirus NS3 protein.
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 beta-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.
authors.affiliation: Structural & Computational Biology Division, School of Biological Sciences, Nanyang Technological University, Singapore.
authors.forename: Dahai
authors.initials: D
authors.lastname: Luo
authors.forename: Ting
authors.initials: T
authors.lastname: Xu
authors.forename: Randall P
authors.initials: RP
authors.lastname: Watson
authors.forename: Daniella
authors.initials: D
authors.lastname: Scherer-Becker
authors.forename: Aruna
authors.initials: A
authors.lastname: Sampath
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Jahnke
authors.forename: Sui Sum
authors.initials: SS
authors.lastname: Yeong
authors.forename: Chern Hoe
authors.initials: CH
authors.lastname: Wang
authors.forename: Siew Pheng
authors.initials: SP
authors.lastname: Lim
authors.forename: Alex
authors.initials: A
authors.lastname: Strongin
authors.forename: Subhash G
authors.initials: SG
authors.lastname: Vasudevan
authors.forename: Julien
authors.initials: J
authors.lastname: Lescar
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism*; D008958:Models, Molecular; D020836:Protein Structure, Quaternary; D012313:RNA / Q000378:metabolism*; D020365:RNA Helicases / Q000737:chemistry* / Q000378:metabolism*; D012697:Serine Endopeptidases / Q000737:chemistry / Q000378:metabolism; D017361:Viral Nonstructural Proteins / Q000737:chemistry* / Q000378:metabolism*; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases
chemical_list: D000255:Adenosine Triphosphate; D012313:RNA; D020365:RNA Helicases; D012697:Serine Endopeptidases; D017361:Viral Nonstructural Proteins; C060028:NS3 protein, flavivirus; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases