Crystal structures of the SARS-CoV-2 nucleocapsid protein C-terminal domain and development of nucleocapsid-targeting...

The ongoing outbreak of COVID-19 caused by SARS-CoV-2 has resulted in a serious public health threat globally. Nucleocapsid protein is a major structural protein of SARS-CoV-2 that plays important roles in the viral RNA packing, replication, assembly, and infection. Here, we report two crystal structures of nucleocapsid protein C-terminal domain (CTD) at resolutions of 2.0 Å and 3.1 Å, respectively. These two structures, crystallized under different conditions, contain 2 and 12 CTDs in asymmetric unit, respectively. Interestingly, despite different crystal packing, both structures show a similar dimeric form as the smallest unit, consistent with its solution form measured by the size-exclusion chromatography, suggesting an important role of CTD in the dimerization of nucleocapsid proteins. By analyzing the surface charge distribution, we identified a stretch of positively charged residues between Lys257 and Arg262 that are involved in RNA-binding. Through screening a single-domain antibodies (sdAbs) library, we identified four sdAbs targeting different regions of nucleocapsid protein with high affinities that have future potential to be used in viral detection and therapeutic purposes.