Mechanistic understanding and inhibition of Zika NS5 protein

Mechanistic understanding and inhibition of Zika NS5 and SARS-CoV-2 RdRP proteinsABSTRACT Zika virus (ZIKV) and Coronavirus (CoV) are single-stranded RNA viruses that pose grave threat to publichealth. In the first two decades of the 21st century, the global community has already witnessed one outbreakof Flavivirus, Zika virus (ZIKV), and three zoonotic outbreaks of CoV- severe acute respiratory syndrome(SARS)-associated coronavirus (SARS-CoV) in 2002, the Middle East respiratory syndrome (MERS)-CoV in2012, and the most recent, the novel SARS-CoV-2. These viruses are highly transmissible, greatly impactingthe social, societal and economic dynamics. However, there are currently no approved drugs for either ZIKV orfor zoonotic CoV, raising an urgent need for development of novel therapeutic strategies against ZIKV andCoV infection. This application seeks to develop an antiviral strategy targeting the viral core replicationmachinery, RNA-dependent RNA polymerase (RdRP), non-structural protein 5 (NS5) of ZIKV and non-structural protein 12 (NSP12) of SARS-CoV-2. On one hand, the currently identified small molecule inhibitorswill be evaluated for their efficiency on ZIKV or SARS-CoV-2 inhibition. On the other hand, mechanistic detailsof ZIKV NS5 and SARS-CoV-2 RdRP-mediated RNA replication will be investigated, thereby providing a basisfor development of synergistic inhibition strategies targeting various enzymatic steps of ZIKV NS5 and SARS-CoV-2. In Aim 1, structural, biochemical and cellular approaches will be taken to evaluate the inhibition of ZIKVNS5- or SARS-CoV-2 RdRP-mediated de novo RNA synthesis by candidate inhibitors. Through evaluation ofthe inhibitory effects of the candidate inhibitors on ZIKV NS5 or SARS-CoV-2 RdRP, this application willaddress whether these compounds can serve as inhibitors to ZIKV NS5 or SARS-CoV-2, and more importantly,to provide a basis for structure-based drug optimization for ZIKV NS5 or SARS-CoV-2. In Aim 2, themechanistic basis of ZIKV NS5 and SARS-CoV-2 RdRP-mediated RNA replication will be determined throughstructure elucidation of the replication complexes of ZIKV NS5 or SARS-CoV-2 RdRP, combined withenzymatic analyses. The structural knowledge on the replication complexes of ZIKV NS5 and SARS-CoV-2RdRP will then provide a framework for structure-based drug design for comprehensive inhibition of ZIKV NS5and SARS-CoV-2 infection. Together, the proposed studies will provide key mechanistic insights into the viralRdRP-mediated genome replication and establish a foundation for development of effective inhibitors againstZIKV and SARS-CoV-2.