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Infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is initiated by virus binding to the ACE2 cell-surface receptors.sup.1-4, followed by fusion of the virus and cell membranes to release the virus genome into the cell. Both receptor binding and membrane fusion activities are mediated by the virus spike glycoprotein.sup.5-7. As with other class-I membrane-fusion proteins, the spike protein is post-translationally cleaved, in this case by furin, into the S1 and S2 components that remain associated after cleavage.sup.8-10. Fusion activation after receptor binding is proposed to involve the exposure of a second proteolytic site (S2'), cleavage of which is required for the release of the fusion peptide.sup.11,12. Here we analyse the binding of ACE2 to the furin-cleaved form of the SARS-CoV-2 spike protein using cryo-electron microscopy. We classify ten different molecular species, including the unbound, closed spike trimer, the fully open ACE2-bound trimer and dissociated monomeric S1 bound to ACE2. The ten structures describe ACE2-binding events that destabilize the spike trimer, progressively opening up, and out, the individual S1 components. The opening process reduces S1 contacts and unshields the trimeric S2 core, priming the protein for fusion activation and dissociation of ACE2-bound S1 monomers. The structures also reveal refolding of an S1 subdomain after ACE2 binding that disrupts interactions with S2, which involves Asp614.sup.13-15 and leads to the destabilization of the structure of S2 proximal to the secondary (S2') cleavage site. Cryo-electron microscopy structures of consecutive binding events of ACE2 in complex with the spike protein of SARS-CoV-2 reveal the mechanisms of receptor binding by the spike protein and activation for membrane fusion by the spike protein of SARS-CoV-2., Author(s): Donald J. Benton [sup.1] , Antoni G. Wrobel [sup.1] , Pengqi Xu [sup.2] [sup.3] , Chloë Roustan [sup.4] , Stephen R. Martin [sup.1] , Peter B. Rosenthal [sup.5] , [...]

Of the 132 people known to have been infected with H7N9 influenza viruses in China, 37 died, and many were severely ill (1). Infection seems to have involved contact with infected poultry (2,3). We have examined the receptor-binding properties of this H7N9 virus and compared them with those of an avian H7N3 virus. We find that the human H7 virus has significantly higher affinity for α-2, 6-linked sialic acid analogues ('human receptor') than avian H7 while retaining the strong binding to α-2,3-linked sialic acid analogues ('avian receptor') characteristic of avian viruses. The human H7 virus does not, therefore, have the preference for human versus avian receptors characteristic of pandemic viruses. X-ray crystallography of the receptor-binding protein, haemagglutinin (HA), in complex with receptor analogues indicates that both human and avian receptors adopt different conformations when bound to human H7 HA than they do when bound to avian H7 HA. Human receptor bound to human H7 HA exits the binding site in a different direction to that seen in complexes formed by HAs from pandemic viruses (4,5) and from an aerosol-transmissible H5 mutant (6). The human-receptor-binding properties of human H7 probably arise from the introduction of two bulky hydrophobic residues by the substitutions Gln226Leu and Gly186Val. The former is shared with the 1957 H2 and 1968 H3 pandemic viruses and with the aerosol-transmissible H5 mutant. We conclude that the human H7 virus has acquired some of the receptor-binding characteristics that are typical of pandemic viruses, but its retained preference for avian receptor may restrict its further evolution towards a virus that could transmit efficiently between humans, perhaps by binding to avian-receptor-rich mucins in the human respiratory tract (7) rather than to cellular receptors., Genetic analyses of the avian H7N9 influenza viruses, responsible for the zoonotic infections of humans in China since February 2013, indicate that the genes for the haemagglutinin (HA) and neuraminidase [...]

Cell-surface-receptor binding by influenza viruses is a key determinant of their transmissibility, both from avian and animal species to humans as well as from human to human. Highly pathogenic avian [...]

Author(s): Shinya Yamada [1, 3]; Yasuo Suzuki [3, 4]; Takashi Suzuki [3, 5]; Mai Q. Le [6]; Chairul A. Nidom [7]; Yuko Sakai-Tagawa [1, 3]; Yukiko Muramoto [1, 3]; Mutsumi [...]

Author(s): Bing Chen [1]; Erik M. Vogan [1, 2]; Haiyun Gong [1]; John J. Skehel [3]; Don C. Wiley [1, 2, 4]; Stephen C. Harrison (corresponding author) [1, 2] The [...]

Key Words mutants, antigenicity, reassortment, recombination, pathogenicity Abstract Influenza A viruses contain genomes composed of eight separate segments of negative-sense RNA. Circulating human strains are notorious for their tendency to [...]

This review describes the three recently established conformational structures of the Hong Kong influenza virus hemagglutinin. Data are discussed with reference to the activity of hemagglutinin in receptor binding, membrane fusion during viral entry, and their implications in the onset of influenza epidemics.

The structure of a stable recombinant ectodomain of influenza hemagglutinin H[A.sub.2] subunit, EH[A.sub.2] (23-185), defined by proteolysis studies of the intact bacterial-expressed ectodomain, was determined to 1.9-[Angstrom] resolution by using x-ray crystallography. The structure reveals a domain composed of N- and C-terminal residues that form an N cap terminating both the N-terminal [Alpha]-helix and the central coiled coil. The N cap is formed by a conserved sequence, and part of it is found in the neutral pH conformation of HA. The C-terminal 23 residues of the ectodomain form a 72-[Angstrom] long nonhelical structure ordered to within 7 residues of the transmembrane anchor. The structure implies that continuous [Alpha] helices are not required for membrane fusion at either the N or C termini. The difference in stability between recombinant molecules with and without the N cap sequences suggests that additional free energy for membrane fusion may become available after the formation of the central triple-stranded coiled coil and insertion of the fusion peptide into the target membrane.

The molecular arrangement of protein complexes engaged in neurotransmitter release and intracellular vesicular flow has been found to be similar to the structures of certain virus membrane fusion proteins. This arrangement is hypothesized to result in close membrane apposition since the complexes are organized for membrane fusion. Results of a study show that complexes from synapses and intracellular vesicles and the membrane fusion subunits of a set of virus glycoproteins create rod-formed alpha-helical bundles. Although they are quite dissimilar in structure, these bundles seem to possess the membrane-anchoring sequences at one of the rod ends.

Research was conducted to examine the hypothesis that a polar octapeptide fused to the N-terminal fusion peptide solubilizes the influenza virus haemagglutinin HA2 subunit ectodomain. The generated chimeric protein F185 folded spontaneously into a soluble trimer with a high alpha-helical content and a high melting temperature, which are the structural characteristics of the low-pH-induced conformation of HA2. Results suggest that the ectodomains of HA2 made in bacteria have membrane-binding properties similar to those of the same ectodomain produced by low-pH treatment of HA isolated from virus.

The ectodomain of the Ebola virus Gp2 glycoprotein was solubilized with a trimeric, isoleucine zipper derived from GCN4 (pIIGCN4) in place of the hydrophobic fusion peptide at the N terminus. This chimeric molecule forms a trimeric, highly [Alpha]-helical, and very thermostable molecule, as determined by chemical crosslinking and circular dichroism. Electron microscopy indicates that Gp2 folds into a rod-like structure like influenza HA2 and HIV-1 gp41, providing further evidence that viral fusion proteins from diverse families such as Orthomyxoviridae (Influenza), Retroviridae (HIV-1), and Filoviridae (Ebola) share common structural features, and suggesting a common membrane fusion mechanism.

A study was conducted to examine the folding of the membrane-anchoring chain of influenza virus hemagglutinin (HA2) in Escherichia coli into the low pH-induced conformation of viral HA2. Crosslinking studies showed that HA2 polypeptide residues 38-175 fold in vivo into a soluble trimer. Crystallography showed that the low pH-induced fold of HA2 is the lowest-energy-state fold of the HA2 polypeptide upon the removal of the receptor-binding chain.

Analysis of the three-dimensional structure of influenza virus hemagglutinin reveals that the secondary and tertiary structures of the molecule refold at low pH, causing a conformational shift that induces merging of host and viral cell membrane. The fusion peptide in the virus relocates by a distance of 100 A to the N-terminal of the molecules. The helical unit at the C-terminal of the molecule unfolds. Relocation of the fusion peptide destabilizes the tripod organization of the molecule and a partial disordering of the structure occurs.