F NMR Reveals multiple conformations at the dimer interface of the nonstructural protein 1 effector domain from influenza A virus

Nonstructural protein 1 of influenza A virus (NS1A) is a conserved virulence factor comprised of an N-terminal double-stranded RNA (dsRNA)-binding domain and a multifunctional C-terminal effector domain (ED), each of which can independently form symmetric homodimers. Here we apply ¹⁹F NMR to NS1A from influenza A/Udorn/307/1972 virus (H3N2) labeled with 5-fluorotryptophan, and we demonstrate that the ¹⁹F signal of Trp187 is a sensitive, direct monitor of the ED helix:helix dimer interface. ¹⁹F relaxation dispersion data reveal the presence of conformational dynamics within this functionally important protein:protein interface, whose rate is more than three orders of magnitude faster than the kinetics of ED dimerization. ¹⁹F NMR also affords direct spectroscopic evidence that Trp187, which mediates intermolecular ED:ED interactions required for cooperative dsRNA binding, is solvent exposed in full-length NS1A at concentrations below aggregation. These results have important implications for the diverse roles of this NS1A epitope during influenza virus infection.