Affinity Purification Mass Spectrometry Analysis of Mutant and Wildtype SARS-CoV-2 Nucleocapsid Protein

The COVID-19 pandemic is arguably the most important global threat to humankind that the world has seen in a century. SARS-CoV-2 has now reached 196 countries across all continents with over 159 million confirmed cases with 3.3 million deaths todate. Currently, our knowledge on how and why SARS-CoV-2 causes severe COVID-19 is continually evolving. Here, we identified three consecutive SNPs (G28881A, G28882A, G28883C) resulting in the Nucleocapsid (N) protein amino acid mutations R203K and G204R (simply termed KR mutation) by large-scale SARS-CoV-2 genome sequencing from clinical swab samples of patients. The nucleocapsid (N) protein of SARS-CoV-2, the highly abundant structure protein within the infected cells, serves multiple functions during viral infection, which besides RNA binding, playing essential roles in viral transcription, replication, and translation. We investigated the impact of this KR mutation on the functional properties of N protein in terms of host protein interaction and phosphorylation status by mass spectrometry.