title: Modulation of the nucleoside triphosphatase/RNA helicase and 5'-RNA triphosphatase activities of Dengue virus type 2 nonstructural protein 3 (NS3) by interaction with NS5, the RNA-dependent RNA polymerase.
abstract: Dengue virus type 2 (DEN2), a member of the Flaviviridae family, is a re-emerging human pathogen of global significance. DEN2 nonstructural protein 3 (NS3) has a serine protease domain (NS3-pro) and requires the hydrophilic domain of NS2B (NS2BH) for activation. NS3 is also an RNA-stimulated nucleoside triphosphatase (NTPase)/RNA helicase and a 5'-RNA triphosphatase (RTPase). In this study the first biochemical and kinetic properties of full-length NS3 (NS3FL)-associated NTPase, RTPase, and RNA helicase are presented. The NS3FL showed an enhanced RNA helicase activity compared with the NS3-pro-minus NS3, which was further enhanced by the presence of the NS2BH (NS2BH-NS3FL). An active protease catalytic triad is not required for the stimulatory effect, suggesting that the overall folding of the N-terminal protease domain contributes to this enhancement. In DEN2-infected mammalian cells, NS3 and NS5, the viral 5'-RNA methyltransferase/polymerase, exist as a complex. Therefore, the effect of NS5 on the NS3 NTPase activity was examined. The results show that NS5 stimulated the NS3 NTPase and RTPase activities. The NS5 stimulation of NS3 NTPase was dose-dependent until an equimolar ratio was reached. Moreover, the conserved motif, 184RKRK, of NS3 played a crucial role in binding to RNA substrate and modulating the NTPase/RNA helicase and RTPase activities of NS3.
authors.affiliation: Department of Microbiology and Immunology, Georgetown University School of Medicine, Washington D. C. 20057, USA.
authors.forename: Changsuek
authors.initials: C
authors.lastname: Yon
authors.forename: Tadahisa
authors.initials: T
authors.lastname: Teramoto
authors.forename: Niklaus
authors.initials: N
authors.lastname: Mueller
authors.forename: Jessica
authors.initials: J
authors.lastname: Phelan
authors.forename: Vannakambadi K
authors.initials: VK
authors.lastname: Ganesh
authors.forename: Krishna H M
authors.initials: KH
authors.lastname: Murthy
authors.forename: R
authors.initials: R
authors.lastname: Padmanabhan
mesh_terms: D017766:Acid Anhydride Hydrolases / Q000378:metabolism; D001665:Binding Sites; D003716:Dengue Virus / Q000201:enzymology*; D007700:Kinetics; D046912:Multiprotein Complexes; D043583:Nucleoside-Triphosphatase / Q000378:metabolism; D020365:RNA Helicases / Q000378:metabolism; D012324:RNA-Dependent RNA Polymerase / Q000378:metabolism; D012697:Serine Endopeptidases / Q000378:metabolism; D017361:Viral Nonstructural Proteins / Q000378:metabolism*; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases
chemical_list: D017766:Acid Anhydride Hydrolases; D046912:Multiprotein Complexes; D043583:Nucleoside-Triphosphatase; D020365:RNA Helicases; D012324:RNA-Dependent RNA Polymerase; D012697:Serine Endopeptidases; D017361:Viral Nonstructural Proteins; C060028:NS3 protein, flavivirus; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases; C096041:NS5 protein, flavivirus; C023090:RNA triphosphatase