title: Efficient replication of a paramyxovirus independent of full zippering of the fusion protein six-helix bundle domain.
abstract: Enveloped viruses such as HIV and members of the paramyxovirus family use metastable, proteinaceous fusion machineries to merge the viral envelope with cellular membranes for infection. A hallmark of the fusogenic glycoproteins of these pathogens is refolding into a thermodynamically highly stable fusion core structure composed of six antiparallel α-helices, and this structure is considered instrumental for pore opening and/or enlargement. Using a paramyxovirus fusion (F) protein, we tested this paradigm by engineering covalently restricted F proteins that are predicted to be unable to close the six-helix bundle core structure fully. Several candidate bonds formed efficiently, resulting in F trimers and higher-order complexes containing covalently linked dimers. The engineered F complexes were incorporated into recombinant virions efficiently and were capable of refolding into a postfusion conformation without temporary or permanent disruption of the disulfide bonds. They efficiently formed fusion pores based on virus replication and quantitative cell-to-cell and virus-to-cell fusion assays. Complementation of these F mutants with a monomeric, fusion-inactive F variant enriched the F oligomers for heterotrimers containing a single disulfide bond, without affecting fusion complementation profiles compared with standard F protein. Our demonstration that complete closure of the fusion core does not drive paramyxovirus entry may aid the design of strategies for inhibiting virus entry.
authors.affiliation: Institute for Biomedical Sciences, Georgia State University, Atlanta, GA 30303; and.
authors.forename: Melinda A
authors.initials: MA
authors.lastname: Brindley
authors.affiliation: Neurovirology Unit, Division of Experimental Clinical Research, Veterinary Public Health, Vetsuisse Faculty, University of Bern, CH-3012 Bern, Switzerland.
authors.forename: Philippe
authors.initials: P
authors.lastname: Plattet
authors.affiliation: Institute for Biomedical Sciences, Georgia State University, Atlanta, GA 30303; and rplemper@gsu.edu.
authors.forename: Richard Karl
authors.initials: RK
authors.lastname: Plemper
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002645:Chickens; D002522:Chlorocebus aethiops; D003545:Cysteine / Q000378:metabolism; D004220:Disulfides / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008459:Measles virus / Q000502:physiology*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000378:metabolism; D015202:Protein Engineering; D055503:Protein Multimerization; D058849:Protein Refolding; D055550:Protein Stability; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011995:Recombination, Genetic / Q000235:genetics; D014760:Viral Fusion Proteins / Q000737:chemistry* / Q000378:metabolism*; D014779:Virus Replication
chemical_list: D004220:Disulfides; D050505:Mutant Proteins; D014760:Viral Fusion Proteins; D003545:Cysteine
keywords: measles virus; membrane fusion