PhD defence : NMR studies of the molecular basis and functions of viral replication

By Maiia Botova (IBS/Protein Dynamics and Flexibility by NMR Group)

In light of the recent COVID-19 pandemic, the issue of developing antiviral drug strategies has become even more urgent. A critical viral target of SARS-CoV-2 is the replication machinery, particularly the nucleocapsid (N) protein and replication-transcription complex (RTC). Phosphorylation of the N protein in its SR-rich region is crucial for continuing the viral cycle. However, the mechanistic details of the process are unknown. This work describes the effect of the N protein phosphorylation by SRPK1/GSK-3/CK1 and PKA kinases on its molecular function. Using NMR spectroscopy, we found that progressive phosphorylation of the N protein alters protein dynamics by disrupting long-range intramolecular contacts. It was also found that a specifically hyperphosphorylated SR-rich region binds to the RNA binding domain of the N protein, abrogating RNA binding. These observations suggest an auto-inhibitory mechanism involving direct competition with RNA.