Honors College Thesis
 

Characterization of the SARS-CoV-2 Nucleocapsid Protein and the Binding to Different Sections of the SARS-CoV-2 Genome

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https://ir.library.oregonstate.edu/concern/honors_college_theses/wp988s45s

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  • The nucleocapsid protein (N protein) of the severe acute respiratory syndrome coronavirus 2 is known to play an essential part of the virus’ functions. The N protein can protect and package the large 29,903 nucleotide genomic RNA whilst also interacting with structural membrane protein M and the spike protein S. The full length protein has a high level of disorder and subsequently has made biophysical analyses difficult. In this paper, we successfully prepare the N protein, its two independent domains, and four fragments of viral genomic RNA. Biophysical techniques of CD, SEC-MALS, and AUC demonstrate that the N protein is partially disordered protein with two independently folded domains, the primary RNA binding domain and dimerization domain, both of which are flanked by disordered linkers. The N protein is a tight dimer with a dimerization constant of 0.4 μM with the ability to facilitate larger oligomers. NMR studies revealed that the dimeric N protein binds to genomic RNA primarily through a specific domain, but also has binding capabilities in the other domain as well. AUC, and SEC-MALS identified a mixture of RNA bound N protein complexes forming as high as 70 dimers of N bound to the viral RNA.
  • Key Words: NMR, SARS-CoV-2, COVID, Nucleocapsid Protein, C-Terminal Domain, N-Terminal Domain, Viral Genomic RNA Binding
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  • This work is funded by a National Science Foundation EAGER MCB 2034446 to Elisar Barbar. We also acknowledge the support of the Oregon State University NMR Facility funded in part by the National Institutes of Health, HEI Grant 1S10OD018518 and by the M.J. Murdock Charitable Trust grant 2014162. Additionally, research funding was awarded from URSA Engage 2018, CURE 2019, & CURE 2020.
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