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bioRxiv · 10.1101/2024.05.09.593331

Structural and functional analyses of SARS-CoV-2 Nsp3 and its specific interactions with the 5' UTR of the viral genome

Lemak, S.·Skarina, T.·Flick, R.·Patel, D. T.·Stogios, P. J.·Savchenko, A.
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Abstract

Non-structural protein 3 (Nsp3) is the largest open reading frame encoded in the SARS-CoV-2 genome, essential for formation of double-membrane vesicles (DMV) wherein viral RNA replication occurs. We conducted an extensive structure-function analysis of Nsp3 and determined the crystal structures of the Ubiquitin-like 1 (Ubl1), Nucleic Acid Binding (NAB), {beta}-coronavirus-Specific Marker ({beta}SM) domains and a sub-region of the Y domain of this protein. We show that the Ubl1, ADP-ribose phosphatase (ADRP), human SARS Unique (HSUD), NAB, and Y domains of Nsp3 bind the 5 UTR of the viral genome and that the Ubl1 and Y domains possess affinity for recognition of this region, suggesting high specificity. The Ubl1-Nucleocapsid (N) protein complex binds the 5 UTR with greater affinity than the individual proteins alone. Our results suggest that multiple domains of Nsp3, particularly Ubl1 and Y, shepherd the 5 UTR of viral genome during translocation through the DMV membrane, priming the Ubl1 domain to load the genome onto N protein.

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Lemak, S., Skarina, T., Flick, R., Patel, D. T., Stogios, P. J., Savchenko, A.. 2024-05-09. Structural and functional analyses of SARS-CoV-2 Nsp3 and its specific interactions with the 5' UTR of the viral genome. https://doi.org/10.1101/2024.05.09.593331

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