title: Mechanism of NS2B-mediated activation of NS3pro in dengue virus: molecular dynamics simulations and bioassays.
abstract: The NS2B cofactor is critical for proteolytic activation of the flavivirus NS3 protease. To elucidate the mechanism involved in NS2B-mediated activation of NS3 protease, molecular dynamic simulation, principal component analysis, molecular docking, mutagenesis, and bioassay studies were carried out on both the dengue virus NS3pro and NS2B-NS3pro systems. The results revealed that the NS2B-NS3pro complex is more rigid than NS3pro alone due to its robust hydrogen bond and hydrophobic interaction networks within the complex. These potent networks lead to remodeling of the secondary and tertiary structures of the protease that facilitates cleavage sequence recognition and binding of substrates. The cofactor is also essential for proper domain motion that contributes to substrate binding. Hence, the NS2B cofactor plays a dual role in enzyme activation by facilitating the refolding of the NS3pro domain as well as being directly involved in substrate binding/interactions. Kinetic analyses indicated for the first time that Glu92 and Asp50 in NS2B and Gln27, Gln35, and Arg54 in NS3pro may provide secondary interaction points for substrate binding. These new insights on the mechanistic contributions of the NS2B cofactor to NS3 activation may be utilized to refine current computer-based search strategies to raise the quality of candidate molecules identified as potent inhibitors against flaviviruses.
authors.affiliation: Centre for Biomedical & Life Sciences, Singapore Polytechnic, Singapore.
authors.forename: Zhili
authors.initials: Z
authors.lastname: Zuo
authors.forename: Oi Wah
authors.initials: OW
authors.lastname: Liew
authors.forename: Gang
authors.initials: G
authors.lastname: Chen
authors.forename: Pek Ching Jenny
authors.initials: PC
authors.lastname: Chong
authors.forename: Siew Hui
authors.initials: SH
authors.lastname: Lee
authors.forename: Kaixian
authors.initials: K
authors.lastname: Chen
authors.forename: Hualiang
authors.initials: H
authors.lastname: Jiang
authors.forename: Chum Mok
authors.initials: CM
authors.lastname: Puah
authors.forename: Weiliang
authors.initials: W
authors.lastname: Zhu
mesh_terms: D000595:Amino Acid Sequence; D003716:Dengue Virus / Q000502:physiology*; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011485:Protein Binding; D025941:Protein Interaction Mapping; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D012697:Serine Endopeptidases / Q000737:chemistry / Q000378:metabolism*; D017361:Viral Nonstructural Proteins / Q000737:chemistry / Q000378:metabolism*; D000085242:Viral Proteases
chemical_list: D012697:Serine Endopeptidases; D017361:Viral Nonstructural Proteins; C080102:NS2B protein, flavivirus; C504449:NS3 protease, dengue virus; D000085242:Viral Proteases