Archives of Virology

, Volume 148, Issue 4, pp 723–744 | Cite as

Characterization of the RNA-binding activity of VP3, a major structural protein of Infectious bursal disease virus

  • G. Kochan
  • D. Gonzalez
  • J. F. Rodriguez

Summary.

Infectious bursal disease virus (IBDV) is the agent of an immune-depressive disease affecting the poultry industry worldwide. Infection of IBDV leads to expression of five mature virus-encoded proteins. Proteolytic processing of the virus-encoded polyprotein generates VP3 which coats the inner surface of the IBDV capsid. In this report, we describe the characterization of the RNA-binding activity of VP3. For these studies, the VP3 coding region was fused to a histidine tag and expressed in insect cells using a recombinant baculovirus. The histidine-tagged VP3 was affinity-purified and used to study its ability to bind RNA molecules using three complementary methods: (i) Northwestern blotting; (ii) binding of VP3 protein-RNA complexes to nitrocellulose membranes; and (iii) electrophoretic mobility shift assays. The results demonstrated that VP3 efficiently bound ssRNA and dsRNA. Under the experimental conditions used in this study, the formation of VP3-RNA complexes did not depend upon the presence of specific RNA sequences. A series of histidine-tagged VP3 deletion mutants spanning the whole VP3 coding region were generated. The use of these mutants revealed that the VP3 RNA-binding domain layed in a highly conserved 69 aa stretch close to the N-terminus of the protein.

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Copyright information

© Springer-Verlag/Wien 2002

Authors and Affiliations

  • G. Kochan
    • 1
  • D. Gonzalez
    • 1
  • J. F. Rodriguez
    • 1
  1. 1.Departamento de Biología Molecular y Celular, Centro Nacional de Biotecnología, CSIC, Campus Universidad Autónoma de Madrid, Madrid, SpainES

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