Structural Basis for a Convergent Immune Response against Ebola Virus

Ebola virus disease is a severe health problem in Africa. Vaccines that display the Zaire ebolavirus glycoprotein spike complex are a prime component for the effort to combat it. The V(H)3-15/V(lambda)1-40-based class of antibodies was recently discovered to be a common response in individuals who received the Ebola virus vaccines. These antibodies display attractive properties, and thus likely contribute to the efficacy of the vaccines. Here, we use cryo-EM to elucidate how three V(H)3-15/V(lambda)1-40 antibodies from different individuals target the virus and found a convergent mechanism against a partially conserved site on the spike complex. Our study rationalizes the selection of the V(H)3-15/V(lambda)1-40 germline genes for specifically targeting this site and highlights Ebolavirus species-specific sequence divergences that may restrict breadth of V(H)3-15/V(lambda)1-40-based humoral response. The results from this study could help develop improved immunization schemes and further enable the design of immunogens that would be efficacious against a broader set of Ebolavirus species

Mapping of the Lassa virus LAMP1 binding site reveals unique determinants not shared by other old world arenaviruses

<div><p>Cell entry of many enveloped viruses occurs by engagement with cellular receptors, followed by internalization into endocytic compartments and pH-induced membrane fusion. A previously unnoticed step of receptor switching was found to be critical during cell entry of two devastating human pathogens: Ebola and Lassa viruses. Our recent studies revealed the functional role of receptor switching to LAMP1 for triggering membrane fusion by Lassa virus and showed the involvement of conserved histidines in this switching, suggesting that other viruses from this family may also switch to LAMP1. However, when we investigated viruses that are genetically close to Lassa virus, we discovered that they cannot bind LAMP1. A crystal structure of the receptor-binding module from Morogoro virus revealed structural differences that allowed mapping of the LAMP1 binding site to a unique set of Lassa residues not shared by other viruses in its family, illustrating a key difference in the cell-entry mechanism of Lassa virus that may contribute to its pathogenicity.</p></div

Data collection and refinement statistics.

<p>Data collection and refinement statistics.</p

Mapping of the LAMP1 binding site on GP1_LASV.

<p>(A) Structural comparison of the loop preceding β5. A yellow dashed line marks an apparent hydrogen bond. (B) Representative images of LAMP1 pull-down experiments by the indicated GP1_LASV-Fc mutants, each one selected from three independent repeats. The panel is composed of three intact membranes as designated by the horizontal lines. (C) Superimposition of the β–hairpins of GP1_LASV and GP1_MORV presented as Cα traces and side-chains only. (D) Ribbon representation of the β–hairpins and their close vicinity in GP1_MORV and GP1_LASV. (E) Surface representation of GP1_LASV. Residues that were mutated and critically affected, weakly affected, or had no affect on LAMP1 binding are colored purple, pink, and blue, respectively, using the same color scheme used in <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1006337#ppat.1006337.g002" target="_blank">Fig 2E</a>. The histidine triad is red. (F) The crystal structure of GP1_LASV (white surface) docked into the EM density of the trimeric spike complex of LASV at pH 5.0 (EMDB: 3292), shown as green mesh at 1.5 σ. The histidine triad is red and all residues identified as important for LAMP1 binding are purple. The crevices between the 3 GP1 subunits are marked with black arrows.</p

GP1_LCMV and GP1_MORV do not interact with LAMP, although GP1_MORV adopts the same global fold as GP1_LASV.

<p>(A) Pull-down assay by the indicated GP1-Fc fusion proteins. The presence of LAMP1 is detected by anti-LAMP1 antibody, and Fc levels are shown using anti-Fc for load control. The pull-down assay was independently repeated three times, and a representative image is shown. (B) SPR analysis of the indicated GP1-Fc analytes. Each analyte was injected at 500 nM over immobilized distal domain of LAMP1. (C) A phylogenetic tree based on the sequences of the GPCs from the indicated OW mammarenaviruses. The scale bar represents a substitution rate of 0.4 per site. (D) Graphical representation of the 16 chains, each colored differently, that make the asymmetric unit. The chains are traced as tubes with a radius proportional to B-factor. N-linked glycans are shown as spheres. (E) Cα traces of all 16 chains superimposed. N-linked glycans are shown with lines. The cysteine residues that make the three disulfide bonds (marked with a ‘D’) are shown with spheres and are labeled, and secondary structure elements are numbered. (F) Multiple sequence alignment of GP1s from the indicated OW mammarenaviruses showing the secondary structure elements as observed in the crystal structure of GP1_MORV. The numbering of the amino acids is based on the sequence of GPC_MORV. Fully conserved residues are highlighted with red background, and partially conserved residues are shown in red. Numbers below the sequences mark the locations and connectivity of the disulfide bridges. The symbol ‘Ψ’ marks the locations of N-linked glycans seen in the crystal structure. We used ESPript [<a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1006337#ppat.1006337.ref021" target="_blank">21</a>] (<a href="http://espript.ibcp.fr/" target="_blank">http://espript.ibcp.fr</a>) for generating this graphical representation.</p

Rational design of universal immunotherapy for TfR1-tropic arenaviruses

International audienceCertain arenaviruses that circulate in rodent populations can cause life-threatening hemor-rhagic fevers when they infect humans. Due to their efficient transmission, arenaviruses pose a severe risk for outbreaks and might be exploited as biological weapons. Effective countermeasures against these viruses are highly desired. Ideally, a single remedy would be effective against many or even all the pathogenic viruses in this family. However, despite the fact that all pathogenic arenaviruses from South America utilize transferrin receptor 1 (TfR1) as a cellular receptor, their viral glycoproteins are highly diversified, impeding efforts to isolate cross-neutralizing antibodies. Here we address this problem using a rational design approach to target TfR1-tropic arenaviruses with high potency and breadth. The pan-reactive molecule is highly effective against all arenaviruses that were tested, offering a universal therapeutic approach. Our design scheme avoids the shortcomings of previous immu-noadhesins and can be used to combat other zoonotic pathogens

Biomolecular Recognition of the Glycan Neoantigen CA19-9 by Distinct Antibodies.

Glycans decorate the cell surface, secreted glycoproteins and glycolipids, and altered glycans are often found in cancers. Despite their high diagnostic and therapeutic potential, however, glycans are polar and flexible molecules that are quite challenging for the development and design of high-affinity binding antibodies. To understand the mechanisms by which glycan neoantigens are specifically recognized by antibodies, we analyze the biomolecular recognition of the tumor-associated carbohydrate antigen CA19-9 by two distinct antibodies using X-ray crystallography. Despite the potential plasticity of glycans and the very different antigen-binding surfaces presented by the antibodies, both structures reveal an essentially identical extended CA19-9 conformer, suggesting that this conformer's stability selects the antibodies. Starting from the bound structure of one of the antibodies, we use the AbLIFT computational algorithm to design a variant with seven core mutations in the variable domain's light-heavy chain interface that exhibits tenfold improved affinity for CA19-9. The results reveal strategies used by antibodies to specifically recognize glycan antigens and show how automated antibody-optimization methods may be used to enhance the clinical potential of existing antibodies