Coronavirus N protein N-terminal domain (NTD) specifically binds the transcriptional regulatory sequence (TRS) and melts TRS-cTRS RNA duplexes.
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All coronaviruses (CoVs), including the causative agent of severe acute respiratory syndrome (SARS), encode a nucleocapsid (N) protein that harbors two independent RNA binding domains of known structure, but poorly characterized RNA binding properties. We show here that the N-terminal domain (NTD) of N protein from mouse hepatitis virus (MHV), a virus most closely related to SARS-CoV, employs aromatic amino acid-nucleobase stacking interactions with a triple adenosine motif to mediate high-affinity binding to single-stranded RNAs containing the transcriptional regulatory sequence (TRS) or its complement (cTRS). Stoichiometric NTD fully unwinds a TRS-cTRS duplex that mimics a transiently formed transcription intermediate in viral subgenomic RNA synthesis. Mutation of the solvent-exposed Y127, positioned on the beta-platform surface of our 1.75 A structure, binds the TRS far less tightly and is severely crippled in its RNA unwinding activity. In contrast, the C-terminal domain (CTD) exhibits no RNA unwinding activity. Viruses harboring Y127A N mutation are strongly selected against and Y127A N does not support an accessory function in MHV replication. We propose that the helix melting activity of the coronavirus N protein NTD plays a critical accessory role in subgenomic RNA synthesis and other processes requiring RNA remodeling.
author list (cited authors)
Grossoehme, N. E., Li, L., Keane, S. C., Liu, P., Dann, C. E., Leibowitz, J. L., & Giedroc, D. P.
complete list of authors
Grossoehme, Nicholas E||Li, Lichun||Keane, Sarah C||Liu, Pinghua||Dann, Charles E||Leibowitz, Julian L||Giedroc, David P