title: A versatile molecular tagging method for targeting proteins to avian reovirus muNS inclusions. Use in protein immobilization and purification.
abstract: BACKGROUND
  Avian reoviruses replicate in viral factories, which are dense cytoplasmic compartments established by protein-protein interactions. The non-structural protein muNS forms the factory scaffold that attracts other viral components in a controlled fashion. To create such a three-dimensional network, muNS uses several different self-interacting domains.
  
  
  RESULTS
  In this study we have devised a strategy to identify muNS regions containing self-interacting domains, based on the capacity of muNS-derived inclusions to recruit muNS fragments. The results revealed that the muNS region consisting of residues 477-542 was recruited with the best efficiency, and this raised the idea of using this fragment as a molecular tag for delivering foreign proteins to muNS inclusions. By combining such tagging system with our previously established method for purifying muNS inclusions from baculovirus-infected insect cells, we have developed a novel protein purification protocol.
  
  
  CONCLUSIONS
  We show that our tagging and inclusion-targeting system can be a simple, versatile and efficient method for immobilizing and purifying active proteins expressed in baculovirus-infected cells. We also demonstrate that muNS inclusions can simultaneously recruit several tagged proteins, a finding which may be used to generate protein complexes and create multiepitope particulate material for immunization purposes.
authors.affiliation: Department of Biochemistry and Molecular Biology, Faculty of Pharmacy and Center for Research in Biological Chemistry and Molecular Materials, University of Santiago de Compostela, Santiago de Compostela, Spain.
authors.forename: Alberto
authors.initials: A
authors.lastname: Brandariz-Nuñez
authors.forename: Rebeca
authors.initials: R
authors.lastname: Menaya-Vargas
authors.forename: Javier
authors.initials: J
authors.lastname: Benavente
authors.forename: Jose
authors.initials: J
authors.lastname: Martinez-Costas
mesh_terms: D000818:Animals; D001665:Binding Sites / Q000235:genetics; D002460:Cell Line; D002478:Cells, Cultured; D002642:Chick Embryo; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D055767:Immobilized Proteins / Q000235:genetics / Q000378:metabolism*; D015151:Immunoblotting; D007181:Inclusion Bodies, Viral / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D030101:Orthoreovirus, Avian / Q000235:genetics / Q000378:metabolism*; D025941:Protein Interaction Mapping / Q000379:methods; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D018411:Spodoptera; D014162:Transfection; D017361:Viral Nonstructural Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*
chemical_list: D055767:Immobilized Proteins; D011993:Recombinant Fusion Proteins; D017361:Viral Nonstructural Proteins; D049452:Green Fluorescent Proteins