title: The A33-dependent incorporation of B5 into extracellular enveloped vaccinia virions is mediated through an interaction between their lumenal domains.
abstract: There are two mechanisms for the incorporation of B5 into the envelope of extracellular virions produced by orthopoxviruses, one that requires A33 and one that does not. We have hypothesized that the A33-dependent mechanism requires a direct interaction between A33 and B5. In this study, chimeric constructs of A33 and B5/B5-green fluorescent protein (GFP) were used to show that the two proteins interact through their lumenal domains and that the coiled-coil domain of B5 is sufficient for an interaction with A33. Furthermore, our experiments reveal that a transmembrane domain, not necessarily its own, is requisite for the lumenal domain of B5 to interact with A33. In contrast, the lumenal domain of A33 is sufficient for interaction with B5. Furthermore, the lumenal domain of A33 is sufficient to restore the proper localization of B5-GFP in infected cells. Taken together, our results demonstrate that the lumenal domains of A33 and B5 interact and that the interaction is required for the incorporation of B5-GFP into extracellular virions, whereas the incorporation of A33 is independent of B5. These results suggest that viral protein incorporation into extracellular virions is an active process requiring specific protein-protein interactions.
authors.affiliation: Department of Microbiology and Immunology, University of Rochester Medical Center, Rochester, New York, USA.
authors.forename: Winnie M
authors.initials: WM
authors.lastname: Chan
authors.forename: Brian M
authors.initials: BM
authors.lastname: Ward
mesh_terms: D006367:HeLa Cells; D006801:Humans; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014615:Vaccinia / Q000235:genetics / Q000378:metabolism*; D014616:Vaccinia virus / Q000235:genetics / Q000378:metabolism*; D014763:Viral Matrix Proteins / Q000235:genetics / Q000378:metabolism*; D014771:Virion / Q000235:genetics / Q000378:metabolism*
chemical_list: C534336:B5 protein, vaccinia virus; C104258:GPA33 protein, human; D008562:Membrane Glycoproteins; D011993:Recombinant Fusion Proteins; D014763:Viral Matrix Proteins