Carboxyl terminus of severe acute respiratory syndrome coronavirus nucleocapsid protein: self-association analysis and nucleic acid binding characterization

Biochemistry. 2006 Oct 3;45(39):11827-35. doi: 10.1021/bi0609319.

Abstract

Coronavirus nucleocapsid (N) protein envelops the genomic RNA to form long helical nucleocapsid during virion assembly. Since N protein oligomerization is usually a crucial step in this process, characterization of such an oligomerization will help in the understanding of the possible mechanisms for nucleocapsid formation. The N protein of severe acute respiratory syndrome coronavirus (SARS-CoV) was recently discovered to self-associate by its carboxyl terminus. In this study, to further address the detailed understanding of the association feature of this C-terminus, its oligomerization was systematically investigated by size exclusion chromatography and chemical cross-linking assays. Our results clearly indicated that the C-terminal domain of SARS-CoV N protein could form not only dimers but also trimers, tetramers, and hexamers. Further analyses against six deletion mutants showed that residues 343-402 were necessary and sufficient for this C-terminus oligomerization. Although this segment contains many charged residues, differences in ionic strength have no effects on its oligomerization, indicating the absence of electrostatic force in SARS-CoV N protein C-terminus self-association. Gel shift assay results revealed that the SARS-CoV N protein C-terminus is also able to associate with nucleic acids and residues 363-382 are the responsible interaction partner, demonstrating that this fragment might involve genomic RNA binding sites. The fact that nucleic acid binding could promote the SARS-CoV N protein C-terminus to form high-order oligomers implies that the oligomeric SARS-CoV N protein probably combines with the viral genomic RNA in triggering long nucleocapsid formation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence / genetics
  • Binding Sites / genetics
  • Chromatography, Gel
  • Coronavirus Nucleocapsid Proteins
  • Dimerization
  • Genome, Viral / physiology
  • Nucleocapsid / chemistry
  • Nucleocapsid / genetics
  • Nucleocapsid / metabolism*
  • Nucleocapsid Proteins / chemistry
  • Nucleocapsid Proteins / genetics
  • Nucleocapsid Proteins / metabolism*
  • Protein Binding / genetics
  • Protein Structure, Tertiary / genetics
  • RNA, Viral / chemistry
  • RNA, Viral / genetics
  • RNA, Viral / metabolism*
  • Sequence Deletion
  • Severe acute respiratory syndrome-related coronavirus / chemistry
  • Severe acute respiratory syndrome-related coronavirus / physiology*
  • Static Electricity
  • Virus Assembly / physiology*

Substances

  • Coronavirus Nucleocapsid Proteins
  • Nucleocapsid Proteins
  • RNA, Viral