Functional and structural characterization of the inter-domain linker region of the dengue virus serotype 4 NS5 protein

• Main Author: Ting, Trinda Anne
• Other Authors: Lim Siew Pheng
• Format: Final Year Project
• Language: English
• Published: 2015
• Subjects: DRNTU::Science::Biological sciences::Microbiology::Virology
• Online Access: http://hdl.handle.net/10356/63146
• Institution: Nanyang Technological University

Dengue (DENV) NS5 is well conserved and contains a 9 amino acid linker that lies between methylase and RNA-dependent RNA polymerase domains. The crystal structure of DENV-3 FL NS5 is the first dengue NS5 to be successfully documented and can be used as a reference for future plans to crystalise the rest of the dengue NS5 proteins and infer possible intra-molecular interactions within the NS5. Further study of the flexible interdomain linker region (IDLR) and its role in the polymerase activity of the DENV-4 NS5 was conducted by mutating different residues into alanine and then measuring the mutants’ de novo (dn) initiation and elongation abilities. DENV infection cell based experiments were also carried out to measure replicative ability of the mutant NS5 proteins. Results showed that firstly, alanine mutations on the IDLR generally reduced in vitro dnI activity but increased elongation ability, indicating that a loss in intramolecular interactions that result in a conformation that favours and upregulates elongation; secondly, effects could enhance in vitro activity but become lethal in the cell. Together, this study allows for a better understanding of the role of IDLR in the function and structure of DENV-4 FL NS5 in the development of its anti-viral agent.