Name
Structural characterization of the infectious bursal disease virus replication machinery
Presenter
Jose Maria Fernandez Palacios, National Center for Biotechnology
Co-Author(s)
José M. Fernández-Palacios, Javier M. Rodríguez, José R. Castón*
Dept. Structure of Macromolecules, Centro Nacional de Biotecnología/CSIC, 28049 Madrid, Spain.
Abstract Category
Structural Biology
Abstract
IBDV is a non-enveloped dsRNA virus that causes severe immunodeficiency in chickens, leading to high mortality rates and significant economic losses in the poultry industry. Unlike other dsRNA viruses, IBDV is characterized by a single-shelled T = 13 capsid, and its genome is organized as ribonucleoprotein complexes (RNPs), which serve as the functional platforms for viral transcription and replication. RNPs are composed of genomic RNA, the nucleoprotein VP3 and the RNA polymerase VP1. Here, we characterized the structure and function of these IBDV replicative complexes.
VP1 is a 100 kDa RNA-dependent RNA polymerase crucial for IBDV replication. Using single-particle cryo-electron microscopy (cryo-EM), we solved the structure of the VP1 molecule, recombinantly expressed from insect cells, in an active functional state. Structural analysis revealed three previously unreported conformational states in which four -helices of the VP1 C-terminal domain are differentially ordered, which could be relevant for the regulation of its enzymatic activity.
VP3 is a multifunctional protein that participates in several steps of the viral cycle. It is required for efficient RNA synthesis, translation and capsid assembly. VP3 is a 30 kDa protein, and has several flexible regions, which make its structural analysis difficult. However, we have purified a stable and active complex of VP1 and VP3 by co-expression in insect cells and purification via affinity and size-exclusion chromatography. VP3 induces the dimerization of VP1, similar to the dimers found at each end of IBDV RNPs. We propose an interaction model that combines our cryo-EM data with Alphafold analysis.
VP1 is a 100 kDa RNA-dependent RNA polymerase crucial for IBDV replication. Using single-particle cryo-electron microscopy (cryo-EM), we solved the structure of the VP1 molecule, recombinantly expressed from insect cells, in an active functional state. Structural analysis revealed three previously unreported conformational states in which four -helices of the VP1 C-terminal domain are differentially ordered, which could be relevant for the regulation of its enzymatic activity.
VP3 is a multifunctional protein that participates in several steps of the viral cycle. It is required for efficient RNA synthesis, translation and capsid assembly. VP3 is a 30 kDa protein, and has several flexible regions, which make its structural analysis difficult. However, we have purified a stable and active complex of VP1 and VP3 by co-expression in insect cells and purification via affinity and size-exclusion chromatography. VP3 induces the dimerization of VP1, similar to the dimers found at each end of IBDV RNPs. We propose an interaction model that combines our cryo-EM data with Alphafold analysis.