Noah McNally

Introduction. Sars-CoV-2 (severe acute respiratory syndrome coronavirus) has become world famous as the causative agent of the COVID-19 disease.¹ According to the World Health Organization, there have been over 346 million confirmed cases of COVID-19 infections worldwide, as of January 2022.² This pandemic has ushered in a plethora of research on the Sars-CoV-2 virus and its mechanisms of pathogenicity, and one of the viral proteins that is of particular importance within the viral life cycle is nsp3 (non-structural protein 3).³ The papain-like protease (PLpro) domain of this protein is necessary for viral replication, as it proteolytically cleaves nsp1, nsp2, and nsp3 from a polyprotein precursor.⁴ These three non-structural proteins, along with nsp4-nsp16 (which are processed by a different protease, 3CLpro) are essential for viral replication to occur.⁴ Additionally, PLpro has been shown to have deubiquitinating and deISGylating activities, and these actions aid the virus in dysregulating the host’s inflammatory response and by inhibiting the host’s antiviral immune response.⁵ Because of this, new treatments are urgently being searched for and developed.⁶ Methods. In order to visualize PLpro’s binding with important ligands such as ISG15, X-ray crystal structures were obtained of a PLpro-mouse ISG15 complex.⁵ Florescence polarization assays were used to assess the activity and specificity of PLpro with various substrates, including ubiquitinated and ISGylated analogs.⁴ Results. Sars-CoV-2 PLpro was confirmed to have deubiquitinating activity and deISGylating activity; however, the specificity for each of these reactions differed than the previously studied PLpro in other Sars-CoV viruses.⁵ The binding pocket for GRL0617 (an inhibitor of PLpro activity) was elucidated, and it was shown that GRL0617 can cause conformational change in PLpro and block ISG15 from reaching the active site of PLpro, thus inhibiting the deISGylation activity of PLpro.⁷ Furthermore, it was discovered that 2-phenylthiopene and several of its derivatives were able to more effectively inhibit PLpro that GRL0617, with IC₅₀ measurements of approximately 500 nm.⁶ The stronger inhibition is attributed to the positive cooperativity of binding to both the Glu167 residue of PLpro and the BL2 groove⁶ Conclusion. Nsp3 and its PLpro domain have been shown to be essential for the replication of the Sars-CoV-2 virus.⁴ The importance of PLpro is also shown by its role in evading the host immune response.⁴ Thus, developing novel treatments is of vital importance, and compounds such as 2-phenylthiopene offer more efficient inhibition through binding to multiple sites on the PLpro domain.⁶

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