Substitution of two residues in the measles virus nucleoprotein results in an impaired self-association.
The nucleoprotein (N) of measles virus encapsidates viral genomic RNA to form a helical nucleocapsid. Its strong self-association is a major hurdle in determining its high-resolution structure using X-ray crystallography. We report the bacterial expression, purification, and characterization of a variant N that has lost its ability to form nucleocapsid-like structures after substitution of two residues by polar residues. Using immunoprecipitation, circular dichroism, and limited proteolysis studies, we show that this nucleoprotein retains a folding similar to wild-type N. Furthermore, the variant N binds the phosphoprotein, indicating that it retains biochemical relevance. We also present evidence indicating that the N-terminus of N lies at the surface of the nucleocapsid. Beyond the identification of one region of N involved in self-association, our results should facilitate structural studies of N using X-ray crystallography.
Karlin
David
D
Architecture et Fonction des Macromolécules Biologiques, UMR 6098 CNRS et Université Aix-Marseilles I et II, ESIL, Campus de Luminy, 13288 Marseilles Cedex 09, France.
Longhi
Sonia
S
Canard
Bruno
B
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001665:Binding Sites; D002942:Circular Dichroism; D008459:Measles virus / Q000378:metabolism*; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D019251:Nucleocapsid / Q000378:metabolism*; D019590:Nucleocapsid Proteins; D009698:Nucleoproteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism; D017354:Point Mutation; D011233:Precipitin Tests; D012367:RNA, Viral / Q000378:metabolism; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*
D019590:Nucleocapsid Proteins; D009698:Nucleoproteins; C055859:P protein, Sendai virus; D010750:Phosphoproteins; D012367:RNA, Viral; D014764:Viral Proteins; C058314:nucleoprotein, Measles virus