Binding of RNA by the Nucleoproteins of Influenza Viruses A and B

Viruses. 2016 Sep 13;8(9):247. doi: 10.3390/v8090247.


This paper describes a biochemical study for making complexes between the nucleoprotein of influenza viruses A and B (A/NP and B/NP) and small RNAs (polyUC RNAs from 5 to 24 nucleotides (nt)), starting from monomeric proteins. We used negative stain electron microscopy, size exclusion chromatography-multi-angle laser light scattering (SEC-MALLS) analysis, and fluorescence anisotropy measurements to show how the NP-RNA complexes evolve. Both proteins make small oligomers with 24-nt RNAs, trimers for A/NP, and dimers, tetramers, and larger complexes for B/NP. With shorter RNAs, the affinities of NP are all in the same range at 50 mM NaCl, showing that the RNAs bind on the same site. The affinity of B/NP for a 24-nt RNA does not change with salt. However, the affinity of A/NP for a 24-nt RNA is lower at 150 and 300 mM NaCl, suggesting that the RNA binds to another site, either on the same protomer or on a neighbour protomer. For our fluorescence anisotropy experiments, we used 6-fluorescein amidite (FAM)-labelled RNAs. By using a (UC)₆-FAM(3') RNA with 150 mM NaCl, we observed an interesting phenomenon that gives macromolecular complexes similar to the ribonucleoprotein particles purified from the viruses.

Keywords: RNA; assembly; influenza virus; nucleoprotein; oligomerization; ribonucleoprotein.

MeSH terms

  • Binding Sites
  • Nucleocapsid Proteins
  • Orthomyxoviridae / physiology*
  • Protein Binding
  • RNA / metabolism*
  • RNA-Binding Proteins / metabolism*
  • Viral Core Proteins / metabolism*
  • Virus Assembly


  • NP protein, Influenza A virus
  • Nucleocapsid Proteins
  • RNA-Binding Proteins
  • Viral Core Proteins
  • nucleoprotein, influenza B virus
  • RNA