Browsing by Author "Trairak Pisikun"

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    Physical and functional interaction of dengue and host protein
    (Mahidol University. Mahidol University Library and Knowledge Center, 2023) Teera Poyomtip; Ponpan Matangkasombut; Siwanon Jirawatnotai; Trairak Pisikun
    Dengue virus (DENV) is the cause of a life-threatening infectious disease affecting around 390 million people annually. Viral non-structural protein 5 (NS5) is known to form ribonucleic acid (RNA) replication complex with host partners and regulate the RNA synthesis. However, the molecular details remain obscure. Therefore, the understanding of the physical and biochemical nature of the interactions plays a pivotal role in viral research and the interactions could open an avenue to develop a new drug target for the dengue patients. In this study, the researcher studied live virus with the epitope tags inserted in viral genome compared to normal virus. The engineered DENV was infective in several human cell lines, and the tags virus showed the same viral kinetic as the parental virus. The tags allowed co-IP of NS5 protein from the infected cells. Utilizing recombinant virus, the researcher identified novel NS5-interacting partners in Huh-7 cells. This result revealed, for the first time, that ribonucleoproteins and fatty acid biosynthesis complexes were in the NS5-interacting network and worked as viral replicase integrators. The study generated NS5 interaction network using tagged virus for the first time. Moreover, dengue NS5 is a phosphor-protein which the exact role of phosphorylation on this protein is still under investigation. The researcher used in silico analysis to screen the potential kinases, which phosphorylate NS5 and the in vitro kinase assay were performed to confirm the specific kinases which were able to phosphorylate NS5 protein. The role of enzyme kinases in dengue viral infection was accessed by inhibitors, an activator and specific siRNAs. The research results indicated that PKC and Akt could act as a restricting mechanism that modulated the DENV replication and repressed the viral outburst in the host cells. Using combined in silico, genetic engineering and biochemical approaches shaded light on how the DENV protein interacted with various host proteins to survive, and could be explored for therapeutic treatment.