Date Published: August 8, 2013
Publisher: Public Library of Science
Author(s): Brett D. Welch, Ping Yuan, Sayantan Bose, Christopher A. Kors, Robert A. Lamb, Theodore S. Jardetzky, Félix A. Rey.
Paramyxoviruses cause a wide variety of human and animal diseases. They infect host cells using the coordinated action of two surface glycoproteins, the receptor binding protein (HN, H, or G) and the fusion protein (F). HN binds sialic acid on host cells (hemagglutinin activity) and hydrolyzes these receptors during viral egress (neuraminidase activity, NA). Additionally, receptor binding is thought to induce a conformational change in HN that subsequently triggers major refolding in homotypic F, resulting in fusion of virus and target cell membranes. HN is an oligomeric type II transmembrane protein with a short cytoplasmic domain and a large ectodomain comprising a long helical stalk and large globular head domain containing the enzymatic functions (NA domain). Extensive biochemical characterization has revealed that HN-stalk residues determine F specificity and activation. However, the F/HN interaction and the mechanisms whereby receptor binding regulates F activation are poorly defined. Recently, a structure of Newcastle disease virus (NDV) HN ectodomain revealed the heads (NA domains) in a “4-heads-down” conformation whereby two of the heads form a symmetrical interaction with two sides of the stalk. The interface includes stalk residues implicated in triggering F, and the heads sterically shield these residues from interaction with F (at least on two sides). Here we report the x-ray crystal structure of parainfluenza virus 5 (PIV5) HN ectodomain in a “2-heads-up/2-heads-down” conformation where two heads (covalent dimers) are in the “down position,” forming a similar interface as observed in the NDV HN ectodomain structure, and two heads are in an “up position.” The structure supports a model in which the heads of HN transition from down to up upon receptor binding thereby releasing steric constraints and facilitating the interaction between critical HN-stalk residues and F.
The Paramyxoviridae are membrane-enveloped negative-sense single-stranded RNA viruses that infect animals and humans often resulting in significant disease and mortality. Most paramyxoviruses enter cells at neutral pH by fusing their envelope with the plasma membrane of a target cell thereby releasing a ribonucleoprotein complex into the cytoplasm. Paramyxovirus fusion is typically mediated by two glycoproteins on the surface of virions: a trimeric fusion protein, F, with type I viral fusion protein characteristics, and a receptor binding protein variously named HN, H, or G depending on the virus and protein functionality .
Here we observe that the PIV5 HN ectodomain can form a head-stalk interaction similar to the arrangement in the structure of the NDV HN , providing further evidence that this conformational state could regulate F activation across different members of the paramyxovirus family. We further observed that the PIV5 HN protein can adopt a hybrid conformation consisting of 2-heads-down and 2-heads-up, with pairs of dimeric heads moving as unified structural units. This hybrid conformational state demonstrates that the PIV5 HN tetramer has inherent flexibility and potential for asymmetry in the head region dimer-of-dimers, consistent with the possibility that sialic acid receptor binding could reorient head domain dimers, exposing the stalk region for F engagement and activation. This hybrid structure also provides a concrete model for the fully helical head-stalk connection in the hypothesized 4-heads-up conformational state.