title: A Structurally Unresolved Head Segment of Defined Length Favors Proper Measles Virus Hemagglutinin Tetramerization and Efficient Membrane Fusion Triggering.
abstract: Paramyxoviruses include several insidious and ubiquitous pathogens of humans and animals, with measles virus (MeV) being a prominent one. The MeV membrane fusion apparatus consists of a receptor binding protein (hemagglutinin [H]) tetramer and a fusion (F) protein trimer. Four globular MeV H heads are connected to a tetrameric stalk through flexible linkers. We sought here to characterize the function of a 17-residue H-head segment proximal to the stalk that was unresolved in all five MeV H-head crystal or cocrystal structures. In particular, we assessed whether its primary sequence and length are critical for proper protein oligomerization and intracellular transport or for membrane fusion triggering. Extensive alanine substitutions had no effect on fusion triggering, suggesting that sequence identity is not critical for this function. Excessive shortening of this segment reduced or completely abrogated fusion trigger function, while length compensation restored it. We then characterized the mechanism of function loss. Mutated H proteins were efficiently transported to the cell surface, but certain alterations enhancing linker flexibility resulted in accumulation of high-molecular-weight H oligomers. Some oligomers had reduced fusion trigger capacity, while others retained this function. Thus, length and rigidity of the unresolved head segment favor proper H tetramerization and counteract interactions between subunits from different tetramers. The structurally unresolved H-head segment, together with the top of the stalk, may act as a leash to provide the right degree of freedom for the heads of individual tetramers to adopt a triggering-permissive conformation while avoiding improper contacts with heads of neighboring tetramers.
  
  
  OBJECTIVE
  Understanding the molecular mechanism of membrane fusion triggering may allow development of new antiviral strategies. The fusion apparatus of paramyxoviruses consists of a receptor binding tetramer and a fusion protein trimer. Structural analyses of the receptor binding hemagglutinin-neuraminidases of certain paramyxoviruses suggest that fusion triggering is preceded by relocation of its head domains, facilitated by flexible linkers. Having noted a structurally unresolved 17-residue segment linking the globular heads to the tetrameric stalk of the measles virus hemagglutinin (H), we asked whether and how it may facilitate membrane fusion triggering. We conclude that, together with the top of the stalk, the flexible linker keeps H heads on a leash long enough to adopt a triggering-permissive conformation but short enough to limit roaming and improper contacts with heads of neighboring tetramers. All morbillivirus H-protein heads appear to be connected to their stalks through a "leash," suggesting a conserved triggering mechanism.
authors.affiliation: Department of Molecular Medicine, Mayo Clinic, and Virology and Gene Therapy Track, Mayo Graduate School, Rochester, Minnesota, USA.
authors.forename: Chanakha K
authors.initials: CK
authors.lastname: Navaratnarajah
authors.affiliation: Department of Molecular Medicine, Mayo Clinic, and Virology and Gene Therapy Track, Mayo Graduate School, Rochester, Minnesota, USA.
authors.forename: Quincy
authors.initials: Q
authors.lastname: Rosemarie
authors.affiliation: Department of Molecular Medicine, Mayo Clinic, and Virology and Gene Therapy Track, Mayo Graduate School, Rochester, Minnesota, USA cattaneo.roberto@mayo.edu.
authors.forename: Roberto
authors.initials: R
authors.lastname: Cattaneo
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D004252:DNA Mutational Analysis; D006389:Hemagglutinins, Viral / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008459:Measles virus / Q000235:genetics / Q000502:physiology*; D050505:Mutant Proteins / Q000235:genetics / Q000378:metabolism; D055503:Protein Multimerization; D021381:Protein Transport; D017384:Sequence Deletion; D053586:Virus Internalization
chemical_list: D006389:Hemagglutinins, Viral; D050505:Mutant Proteins; C032397:hemagglutinin protein G, measles virus