Purification of Dengue and Zika Virus Non-structural Protein 5 for Crystallization and Screening of Antivirals

Abstract

Dengue Virus (DENV) and ZIKA Virus (ZIKV) are two important human pathogens that belong to the Flavivirus genus of positive strand RNA viruses. Symptoms of DENV infections range from asymptomatic or mild fever to life-threatening forms, while ZIKV can lead to teratogenic effects such as microcephaly in newborns and neurological disease like the Guillain–Barré syndrome. Non-Structural Protein 5 (NS5) is the largest and most conserved enzyme across flaviviruses and hence constitutes a prime target for developing pan-flavivirus antiviral inhibitors. NS5 results from the gene fusion between a methyltransferase at the N-terminus of the protein and an RNA-dependent RNA polymerase (RdRp) at the C-terminal end. The NS5 protein plays key roles in replication and modification of viral RNA and its inhibition by potent antiviral drugs could prevent severe symptoms associated with infections. We have optimized purification and crystallization protocols to obtain active recombinant proteins suitable for structure-based drug discovery for both the full-length NS5 protein and the polymerase domain of NS5 from DENV and ZIKV.