Structure of the nucleoprotein binding domain of Mokola virus phosphoprotein

J Virol. 2010 Jan;84(2):1089-96. doi: 10.1128/JVI.01520-09. Epub 2009 Nov 11.

Abstract

Mokola virus (MOKV) is a nonsegmented, negative-sense RNA virus that belongs to the Lyssavirus genus and Rhabdoviridae family. MOKV phosphoprotein P is an essential component of the replication and transcription complex and acts as a cofactor for the viral RNA-dependent RNA polymerase. P recruits the viral polymerase to the nucleoprotein-bound viral RNA (N-RNA) via an interaction between its C-terminal domain and the N-RNA complex. Here we present a structure for this domain of MOKV P, obtained by expression of full-length P in Escherichia coli, which was subsequently truncated during crystallization. The structure has a high degree of homology with P of rabies virus, another member of Lyssavirus genus, and to a lesser degree with P of vesicular stomatitis virus (VSV), a member of the related Vesiculovirus genus. In addition, analysis of the crystal packing of this domain reveals a potential binding site for the nucleoprotein N. Using both site-directed mutagenesis and yeast two-hybrid experiments to measure P-N interaction, we have determined the relative roles of key amino acids involved in this interaction to map the region of P that binds N. This analysis also reveals a structural relationship between the N-RNA binding domain of the P proteins of the Rhabdoviridae and the Paramyxoviridae.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Binding Sites
  • Crystallization
  • Crystallography, X-Ray
  • Escherichia coli / genetics
  • Escherichia coli / metabolism
  • Lyssavirus / genetics
  • Lyssavirus / metabolism*
  • Models, Molecular
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Nucleocapsid Proteins / chemistry
  • Nucleocapsid Proteins / genetics
  • Nucleocapsid Proteins / metabolism*
  • Phosphoproteins / chemistry*
  • Phosphoproteins / genetics
  • Phosphoproteins / metabolism
  • Structure-Activity Relationship
  • Two-Hybrid System Techniques
  • Viral Proteins / chemistry*
  • Viral Proteins / genetics
  • Viral Proteins / metabolism

Substances

  • Nucleocapsid Proteins
  • Phosphoproteins
  • Viral Proteins

Associated data

  • PDB/2WZL