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Author: Carravilla, Pablo × Author: Eggeling, Christian × End date: 2024 × Start date: 2020 × DDC: 570 × Has files: Yes ×

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    Functional Delineation of a Protein–Membrane Interaction Hotspot Site on the HIV-1 Neutralizing Antibody 10E8
    (Basel : Molecular Diversity Preservation International (MDPI), 2022) Insausti, Sara; Garcia-Porras, Miguel; Torralba, Johana; Morillo, Izaskun; Ramos-Caballero, Ander; de la Arada, Igor; Apellaniz, Beatriz; Caaveiro, Jose M. M.; Carravilla, Pablo; Eggeling, Christian; Rujas, Edurne; Nieva, Jose L.
    Antibody engagement with the membrane-proximal external region (MPER) of the envelope glycoprotein (Env) of HIV-1 constitutes a distinctive molecular recognition phenomenon, the full appreciation of which is crucial for understanding the mechanisms that underlie the broad neutralization of the virus. Recognition of the HIV-1 Env antigen seems to depend on two specific features developed by antibodies with MPER specificity: (i) a large cavity at the antigen-binding site that holds the epitope amphipathic helix; and (ii) a membrane-accommodating Fab surface that engages with viral phospholipids. Thus, besides the main Fab–peptide interaction, molecular recognition of MPER depends on semi-specific (electrostatic and hydrophobic) interactions with membranes and, reportedly, on specific binding to the phospholipid head groups. Here, based on available cryo-EM structures of Fab–Env complexes of the anti-MPER antibody 10E8, we sought to delineate the functional antibody–membrane interface using as the defining criterion the neutralization potency and binding affinity improvements induced by Arg substitutions. This rational, Arg-based mutagenesis strategy revealed the position-dependent contribution of electrostatic interactions upon inclusion of Arg-s at the CDR1, CDR2 or FR3 of the Fab light chain. Moreover, the contribution of the most effective Arg-s increased the potency enhancement induced by inclusion of a hydrophobic-at-interface Phe at position 100c of the heavy chain CDR3. In combination, the potency and affinity improvements by Arg residues delineated a protein–membrane interaction site, whose surface and position support a possible mechanism of action for 10E8-induced neutralization. Functional delineation of membrane-interacting patches could open new lines of research to optimize antibodies of therapeutic interest that target integral membrane epitopes.
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    Affinity for the Interface Underpins Potency of Antibodies Operating In Membrane Environments
    (Maryland Heights, MO : Cell Press, 2020) Rujas, Edurne; Insausti, Sara; Leaman, Daniel P.; Carravilla, Pablo; González-Resines, Saul; Monceaux, Valérie; Sánchez-Eugenia, Rubén; Garcıá-Porras, Miguel; Iloro, Ibon; Zhang, Lei; Elortza, Félix; Julien, Jean-Philippe; Saéz-Cirión, Asier; Zwick, Michael B.; Eggeling, Christian; Ojida, Akio; Domene, Carmen; Caaveiro, Jose M.M.; Nieva, José L.
    The contribution of membrane interfacial interactions to recognition of membrane-embedded antigens by antibodies is currently unclear. This report demonstrates the optimization of this type of antibodies via chemical modification of regions near the membrane but not directly involved in the recognition of the epitope. Using the HIV-1 antibody 10E8 as a model, linear and polycyclic synthetic aromatic compounds are introduced at selected sites. Molecular dynamics simulations predict the favorable interactions of these synthetic compounds with the viral lipid membrane, where the epitope of the HIV-1 glycoprotein Env is located. Chemical modification of 10E8 with aromatic acetamides facilitates the productive and specific recognition of the native antigen, partially buried in the crowded environment of the viral membrane, resulting in a dramatic increase of its capacity to block viral infection. These observations support the harnessing of interfacial affinity through site-selective chemical modification to optimize the function of antibodies that target membrane-proximal epitopes. © 2020 The Author(s)Rujas et al. describe the site-selective chemical modification of antibodies to improve the molecular recognition of epitopes at membrane surfaces. The modification using aromatic compounds dramatically enhanced the virus neutralization potency and native antigen binding efficiency of HIV-1 antibodies directed against the membrane-embedded MPER epitope. © 2020 The Author(s)