Paramyxoviruses (measles, mumps, Nipah, human parainfluenza, Newcastle disease viruses) enter cells through the concerted action of a trimeric fusion glycoprotein (F) and a tetrameric attachment glycoprotein (HN, H, or G). A 2014 study by Bose, Song, Jardetzky, and Lamb consolidated evidence for a conserved core mechanism — the stalk exposure model, first proposed for parainfluenza virus 5 — in which receptor binding by the attachment protein's globular heads exposes a four-helix-bundle (4HB) stalk domain that triggers F refolding, with headless stalk constructs able to trigger F without receptor binding. That work left open the atomic details of the F–HN/H/G interaction, the ordered sequence of conformational changes from receptor binding to F activation, the structural determinants of why only certain stalk lengths are competent, and whether the mechanism is shared across all paramyxovirus genera. A decade of follow-up has constrained the model functionally (stalk-exposure evidence in morbilliviruses and henipaviruses, linker-flexibility requirements, bidentate henipavirus F–G contacts) and structurally (subnanometer cryo-electron tomography of the human parainfluenza virus 3 HN/F complex on authentic virions showing an HN loop inserted into a pocket on F, with one head contacting F and the other reaching for receptor; atomic structures of prefusion F alone). However, no atomic-resolution co-structure of F with any attachment protein exists for any paramyxovirus in any functional state, the active-state stalk rearrangement has not been directly observed, and the henipavirus and morbillivirus F–attachment interfaces remain structurally unresolved. The open core is the atomic-level F–HN/H/G interface and the complete receptor-to-F conformational cascade, including the structural basis of stalk-length dependence and the generality of the triggering architecture across paramyxovirus genera.

Background

Paramyxoviruses are enveloped, negative-sense single-stranded RNA viruses whose members include major human and animal pathogens (measles virus, mumps virus, human parainfluenza viruses, Nipah and Hendra viruses, Newcastle disease virus). Unlike viruses such as influenza or HIV, which use one glycoprotein for both receptor binding and membrane fusion, every paramyxovirus uses two envelope glycoproteins operating at neutral pH: an attachment protein — called HN (hemagglutinin-neuraminidase) in respiro-, rubula-, and avulaviruses, H (hemagglutinin) in morbilliviruses, and G in henipaviruses — and a fusion protein, F. F is a class I viral fusion protein: it folds into a metastable prefusion trimer, is activated by proteolytic cleavage during trafficking, and, once triggered, refolds irreversibly through a prehairpin intermediate into a postfusion six-helix bundle that merges the viral and target-cell membranes. The attachment protein is a disulfide-linked tetramer of dimers comprising a membrane-proximal stalk — structurally a four-helix bundle (4HB), a parallel bundle of four alpha-helices, one from each chain — topped by globular receptor-binding heads.

The source study (Bose, Song, Jardetzky, and Lamb, Journal of Virology, 2014) asked how the attachment protein triggers F. Mutating solvent-exposed hydrophobic stalk residues (PIV5 HN V81, L85, P84; mumps virus HN V98, L102) abolished F activation without affecting surface expression, receptor binding, or neuraminidase activity, defining an F-activating surface on the stalk 4HB. An engineered disulfide (S92C) that rigidified the Newcastle disease virus (NDV) HN stalk blocked fusion, and this block was partially rescued by a hyperfusogenic, destabilized F mutant (L289A) — implicating stalk flexibility and F metastability in the triggering threshold. Most strikingly, headless stalk constructs lacking the entire receptor-binding head (NDV HN 1-123; mumps virus HN 1-132) still triggered F-dependent cell-cell fusion without any receptor binding event, and only the longest mumps HN construct (1-132, not 1-119, 1-122, 1-125, or 1-128) was active, showing that F activation requires a precise stalk length and conformation. Together with parallel measles-virus H and Nipah-virus G studies, these results supported the 'stalk exposure model': in the resting state the globular heads shield the F-activating stalk surface; receptor binding rearranges the heads, exposing the stalk 4HB, which then engages F in an 'induced fit' interaction that lowers F's activation barrier. The central unresolved issues the authors identified were the atomic details of the F–HN/H/G interface, the ordered sequence of conformational changes upon receptor binding, the structural determinants of stalk-length dependence, and whether the mumps HN E105 residue (a charged specificity determinant whose mutation to leucine abolishes activation of heterotypic PIV5 F but not mumps F) functions identically in the isolated stalk; the authors concluded that further investigation into the F-triggering process is necessary in order to confirm the proposed model.

Problem Statement

Determine the atomic structure of the paramyxovirus F–attachment protein (HN/H/G) complex and the mechanistic sequence linking receptor binding to F activation. Specifically: (1) an atomic-resolution structure of F in complex with its attachment protein — stalk and heads — in the pre-trigger, quiescent state, and in the receptor-bound state that exposes the stalk 4HB and triggers F, identifying the residues and geometry of the F-activating interface (including the role of stalk hydrophobic residues and charged specificity determinants such as mumps HN E105); (2) the ordered sequence of conformational changes in the attachment protein (head rearrangement, stalk exposure, induced-fit engagement of F) and in F (prefusion to prehairpin to postfusion) that constitutes the triggering cascade; (3) the structural basis for the strict dependence of F activation on stalk 4HB length and conformation (why mumps HN headless constructs 1-119 through 1-128 are inactive while 1-132 is active); and (4) whether the triggering architecture — the quaternary arrangement and the interface residues by which the attachment protein engages and triggers F — is conserved across paramyxovirus genera — comparing sialic-acid-receptor HN viruses (respiro-, rubula-, avulaviruses) with protein-receptor morbillivirus (H–F) and henipavirus (G–F) systems, for which even the existence of a stable preassembled F–attachment complex is contested.

Because no atomic-resolution co-structure of F with an attachment protein has yet been determined for any paramyxovirus in any functional state, a valid solution is an atomically explicit, mechanistically grounded model rather than newly measured coordinates: it must specify a proposed interface (the residues and geometry of the F–attachment contacts) and an ordered conformational cascade, constrained by the existing structural and functional evidence, and stated so that it makes discriminating, testable predictions. The four items above are jointly required: a solution that resolves only some of them does not resolve the task. Each item is discharged by a stated artifact: (1) by the proposed interface (residues and geometry), (2) by the ordered conformational cascade, (3) by an atomically explicit length-and-conformation rationale that relates the inactive 1-119 through 1-128 constructs to the active 1-132 construct through that interface and cascade, and (4) by an explicit per-genus comparison of the interface residues and ordered cascade across the sialic-acid-receptor HN viruses and the protein-receptor morbillivirus and henipavirus systems.

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