Structure of the Newcastle disease virus hemagglutinin-neuraminidase (HN) ectodomain reveals a four-helix bundle stalk

Abstract

The paramyxovirus hemagglutinin-neuraminidase (HN) protein plays multiple roles in viral entry and egress, including binding to sialic acid receptors, activating the fusion (F) protein to activate membrane fusion and viral entry, and cleaving sialic acid from carbohydrate chains. HN is an oligomeric integral membrane protein consisting of an N-terminal transmembrane domain, a stalk region, and an enzymatically active neuraminidase (NA) domain. Structures of the HN NA domains have been solved previously; however, the structure of the stalk region has remained elusive. The stalk region contains specificity determinants for F interactions and activation, underlying the requirement for homotypic F and HN interactions in viral entry. Mutations of the Newcastle disease virus HN stalk region have been shown to affect both F activation and NA activities, but a structural basis for understanding these dual affects on HN functions has been lacking. Here, we report the structure of the Newcastle disease virus HN ectodomain, revealing dimers of NA domain dimers flanking the N-terminal stalk domain. The stalk forms a parallel tetrameric coiled-coil bundle (4HB) that allows classification of extensive mutational data, providing insight into the functional roles of the stalk region. Mutations that affect both F activation and NA activities map predominantly to the 4HB hydrophobic core, whereas mutations that affect only F-protein activation map primarily to the 4HB surface. Two of four NA domains interact with the 4HB stalk, and residues at this interface in both the stalk and NA domain have been implicated in HN function.

Roles of the highly conserved amino acids in the globular head and stalk region of the Newcastle disease virus HN protein in the membrane fusion process

“…The virus increased neuraminidase activity to promote virus release when its hemagglutinin ability was improved. In the same mutant, the impact on receptor-binding ability, neuraminidase activity, and cell fusion is different (36). The mutations in NDV HN demonstrated that R174L, R416L, R498L, E547Q, Y526L, and I175E impacted the receptor binding ability and NA activity.…”

Identification of a Region in the Stalk Domain of the Nipah Virus Receptor Binding Protein That Is Critical for Fusion Activation

“…The stalk domain structures of NDV (63) and PIV5 (64) have recently been solved, and our chimeric proteins were modeled based on the NDV structure (Fig. 16).…”

Immobilization of the N-terminal helix stabilizes prefusion paramyxovirus fusion proteins

“…4). Together with biochemical and structural data of paramyxovirus attachment proteins (17)(18)(19)(20)(21)(22)(23)(24)(25)(26)(27)(28), a general model of paramyxovirus fusion has been proposed: upon activation by the attachment protein, metastable prefusion F undergoes a series of large-scale, ATP-independent conformational changes, going down an energy gradient from a metastable prefusion state to a highly stable postfusion state. The energy released during F refolding is believed to facilitate membrane fusion to create a pore between the virus and host cell through which the viral ribonucleoprotein complex can enter the target cell.…”