Crystal Structure of the Pre-fusion Nipah Virus Fusion Glycoprotein Reveals a Novel Hexamer-of-Trimers Assembly

Xu, Kai; Chan, Yee Peng; Bradel-Tretheway, Birgit; Akyol-Ataman, Zeynep; Zhu, Yongqun; Dutta, Somnath; Yan, Lianying; Yan, Feng; Wang, Lin‐Fa; Skiniotis, Georgios; Lee, Benhur; Zhou, Z. Hong; Broder, Christopher C.; Aguilar, Hector C.; Nikolov, Dimitar B.

doi:10.1371/journal.ppat.1005322

Cited by 72 publications

(91 citation statements)

References 56 publications

(56 reference statements)

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“…Targeted deletion regions were chosen based on X-ray crystallographic images for NiV and other paramyxoviruses (45)(46)(47)(48), and protein regions were chosen that appeared to be exposed and therefore had the potential to interact with their partner glycoprotein or that looped out, suggesting that they could potentially be removed without destroying the overall integrity of the glycoprotein.…”

Section: Identification Of Dispensable Regions For G-f Interactionsmentioning

confidence: 99%

Multiple Strategies Reveal a Bidentate Interaction between the Nipah Virus Attachment and Fusion Glycoproteins

Stone

Vemulapati

Bradel-Tretheway

et al. 2016

J Virol

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The paramyxoviral family contains many medically important viruses, including measles virus, mumps virus, parainfluenza viruses, respiratory syncytial virus, human metapneumovirus, and the deadly zoonotic henipaviruses Hendra and Nipah virus (NiV). To both enter host cells and spread from cell to cell within infected hosts, the vast majority of paramyxoviruses utilize two viral envelope glycoproteins: the attachment glycoprotein (G, H, or hemagglutinin-neuraminidase [HN]) and the fusion glycoprotein (F). Binding of G/H/HN to a host cell receptor triggers structural changes in G/H/HN that in turn trigger F to undergo a series of conformational changes that result in virus-cell (viral entry) or cell-cell (syncytium formation) membrane fusion. The actual regions of G/H/HN and F that interact during the membrane fusion process remain relatively unknown though it is generally thought that the paramyxoviral G/H/HN stalk region interacts with the F head region. Studies to determine such interactive regions have relied heavily on coimmunoprecipitation approaches, whose limitations include the use of detergents and the micelle-mediated association of proteins. Here, we developed a flow-cytometric strategy capable of detecting membrane proteinprotein interactions by interchangeably using the full-length form of G and a soluble form of F, or vice versa. Using both coimmunoprecipitation and flow-cytometric strategies, we found a bidentate interaction between NiV G and F, where both the stalk and head regions of NiV G interact with F. This is a new structural-biological finding for the paramyxoviruses. Additionally, our studies disclosed regions of the NiV G and F glycoproteins dispensable for the G and F interactions. IMPORTANCENipah virus (NiV) is a zoonotic paramyxovirus that causes high mortality rates in humans, with no approved treatment or vaccine available for human use. Viral entry into host cells relies on two viral envelope glycoproteins: the attachment (G) and fusion (F) glycoproteins. Binding of G to the ephrinB2 or ephrinB3 cell receptors triggers conformational changes in G that in turn cause F to undergo conformational changes that result in virus-host cell membrane fusion and viral entry. It is currently unknown, however, which specific regions of G and F interact during membrane fusion. Past efforts to determine the interacting regions have relied mainly on coimmunoprecipitation, a technique with some pitfalls. We developed a flow-cytometric assay to study membrane protein-protein interactions, and using this assay we report a bidentate interaction whereby both the head and stalk regions of NiV G interact with NiV F, a new finding for the paramyxovirus family.T he paramyxovirus family includes many important negativesense, single-stranded enveloped RNA viruses, such as measles (MeV), mumps (MuV), respiratory syncytial (RSV), Newcastle disease (NDV), parainfluenza (PIV) Nipah (NiV), and Hendra (HeV) viruses and human metapneumovirus (hMPV) (1). With very few exceptions, paramyxoviruses require two di...

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Section: Identification Of Dispensable Regions For G-f Interactionsmentioning

confidence: 99%

Multiple Strategies Reveal a Bidentate Interaction between the Nipah Virus Attachment and Fusion Glycoproteins

Stone

Vemulapati

Bradel-Tretheway

et al. 2016

J Virol

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“…While electron microscopy has established a basic framework for the paramyxovirus virion organization [3,4,5,6,7,8,9,10,11,12,13,14,15,16,17,18,19,20,21,22,23] and a number of crystal structures of paramyxovirus proteins and protein fragments have been solved (for instance [6,24,25,26,27,28,29,30,31,32,33,34,35,36,37,38,39,40,41,42], the reconstruction of the 3D ultrastructures of paramyxovirus particles is impaired by particle size and the pleomorphic nature of the virions, which prevents single particle reconstruction approaches. To overcome the problem, recent studies have applied cryo-electron tomography (cryo-ET) to the analysis of paramyxovirus particles.…”

Section: Introductionmentioning

confidence: 99%

Structure and organization of paramyxovirus particles

Cox

Plemper

2017

Current Opinion in Virology

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The paramyxovirus family comprises major human and animal pathogens such as measles virus (MeV), mumps virus (MuV), the parainfluenzaviruses, Newcastle disease virus (NDV), and the highly pathogenic zoonotic hendra (HeV) and nipah (NiV) viruses. Paramyxovirus particles are pleomorphic, with a lipid envelope, nonsegmented RNA genomes of negative polarity, and densely packed glycoproteins on the virion surface. A number of crystal structures of different paramyxovirus proteins and protein fragments were solved, but the available information concerning overall virion organization remains limited. However, recent studies have reported cryo-electron tomography-based reconstructions of Sendai virus (SeV), MeV, NDV, and human parainfluenza virus type 3 (HPIV3) particles and a surface assessment of NiV-derived virus-like particles (VLPs), which have yielded innovative hypotheses concerning paramyxovirus particle assembly, budding, and organization. Following a summary of the current insight into paramyxovirus virion morphology, this review will focus on discussing the implications of these particle reconstructions on the present models of paramyxovirus assembly and infection.

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“…Distance and contact measurements were calculated using VMD plugins. For distance measurements between the N-terminal helix and the hydrophobic pocket, α-carbon coordinates of residue 22 and residue 287 were used, where residue 287 was determined to be the center of mass of the hydrophobic pocket, which remains well-defined throughout the simulation as well as among prefusion and postfusion structures (9)(10)(11)(12)(13)(14)(15)(16). For hydrophobic contact measurements, a cutoff distance of 4 Å between carbon atoms was used.…”

Section: Methodsmentioning

confidence: 99%

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

Song¹,

Poor²,

Abriata

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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Parainfluenza virus 5 (PIV5) is an enveloped, single-stranded, negative-sense RNA virus of the Paramyxoviridae family. PIV5 fusion and entry are mediated by the coordinated action of the receptor-binding protein, hemagglutinin–neuraminidase (HN), and the fusion protein (F). Upon triggering by HN, F undergoes an irreversible ATP- and pH-independent conformational change, going down an energy gradient from a metastable prefusion state to a highly stable postfusion state. Previous studies have highlighted key conformational changes in the F-protein refolding pathway, but a detailed understanding of prefusion F-protein metastability remains elusive. Here, using two previously described F-protein mutations (S443D or P22L), we examine the capacity to modulate PIV5 F stability and the mechanisms by which these point mutants act. The S443D mutation destabilizes prefusion F proteins by disrupting a hydrogen bond network at the base of the F-protein globular head. The introduction of a P22L mutation robustly rescues destabilized F proteins through a local hydrophobic interaction between the N-terminal helix and a hydrophobic pocket. Prefusion stabilization conferred by a P22L-homologous mutation is demonstrated in the F protein of Newcastle disease virus, a paramyxovirus of a different genus, suggesting a conserved stabilizing structural element within the paramyxovirus family. Taken together, the available data suggest that movement of the N-terminal helix is a necessary early step for paramyxovirus F-protein refolding and presents a novel target for structure-based drug design.

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Crystal Structure of the Pre-fusion Nipah Virus Fusion Glycoprotein Reveals a Novel Hexamer-of-Trimers Assembly

Cited by 72 publications

References 56 publications

Multiple Strategies Reveal a Bidentate Interaction between the Nipah Virus Attachment and Fusion Glycoproteins

Multiple Strategies Reveal a Bidentate Interaction between the Nipah Virus Attachment and Fusion Glycoproteins

Structure and organization of paramyxovirus particles

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

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