Search bioRxiv⌕ Search

Biology subjects

Hastie, K.

Publications and source records attributed to Hastie, K..

4 recordsLinked to original sources

Structural and Mechanistic Basis for Antibody Neutralization of the Measles Fusion Protein

Measles virus (MeV) is a highly contagious viral pathogen and remains a major global health threat. Resurgent infections, driven by insufficient vaccine coverage, waning herd immunity, and the vulnerability of immunocompromised individuals, highlight the urgent need for effective countermeasures. Monoclonal antibodies (mAbs) represent a promising strategy, both as antiviral agents and as probes of viral entry mechanisms. While most vaccine-elicited neutralizing antibodies target the hemagglutinin (H) protein, emerging evidence suggests that antibodies against the fusion (F) protein are also potent inhibitors. Still, there is insufficient information on the target sites and activities of antibodies against the F protein. Like other class I fusion proteins, MeV F exists in a metastable prefusion state that undergoes dramatic conformational changes during viral entry. Here, we selected four mAbs that recognize conformational patterns of F-prefusion and/or postfusion, characterized their epitopes, specificities, and antiviral activities. Structural analyses mapped antibody interactions onto pre- and postfusion F conformations, revealing that all three neutralizing mAbs are specific for the prefusion form, while the non-neutralizing mAb recognizes only the postfusion F. Biophysical and functional assays defined distinct mechanisms: neutralization occurs either by stabilizing the prefusion protein or by preventing the extended intermediate from completing fusion. We also describe a novel mechanism of neutralization in which an antibody prematurely triggers F activation but blocks the subsequent refolding required for viral entry. Together, these findings provide the first detailed mapping of neutralizing epitopes on the MeV F protein and establish a framework for the rational design of F-targeted intervention.

immunology↗

Uncovering the features of Measles-targeting human antibodies elicited by the MMR vaccine

Measles virus (MeV), a highly transmissible paramyxovirus, causes disease that can lead to severe complications and death, particularly in babies and young children. Deployment of the durable, highly effective, live-attenuated measles vaccine has saved an estimated 94 million lives in the past 50 years,1 yet the immunological explanation for this vaccines unique success and its landscape of antibody recognition remains unclear. Here we report the first panel of human monoclonal antibodies (mAbs) specific for the MeV hemagglutinin (H) and fusion (F) surface proteins, derived from the memory B cells of an MMR vaccinee. From over 100 cloned human mAbs, we mapped four major epitope clusters on H and another five major clusters on F, and structurally characterized 17 representative mAbs including one or more examples of each of the nine epitope clusters on the two surface antigens. We find that antibodies against both H and F can lead to potent virus neutralization and reduction of viral loads in vivo, including one mAb against F that reduces viral loads to below the limit of detection for all animals. High-resolution cryo-EM reveals contact sites of the most protective antibodies against both surface antigens. Discovery, characterization, and in vitro and in vivo success of these fully human mAbs now provide new avenues for prophylactic or therapeutic intervention against this re-emerging virus. HighlightsO_LIA large panel of Measles-specific monoclonal antibodies was isolated from a human MMR vaccinee, years after vaccination. C_LIO_LIStructural and biochemical mapping paints a landscape of antibody recognition with nine major competition groups, including four major sites on Hemagglutinin (H) and five on the fusion protein (F). C_LIO_LIAntibodies against both H and F confer in vitro neutralization and in vivo protection, including, in one case, undetectable viral load after antibody treatment. C_LIO_LIThe most protective H-specific mAbs, 4D08 and 1C08, target the receptor-binding site and the F-proximal outside of the H dimer, and likely function by interfering with receptor binding and H-F interactions, respectively. C_LIO_LIThe most protective F-specific mAbs, 3A12 and 4F09, target the sides and apex of the prefusion F trimer, and likely function by locking F into its pre-fusion state C_LI

immunology↗

A broadly-neutralizing antibody against Ebolavirus glycoprotein that potentiates the breadth and neutralization potency of other antibodies

Ebolavirus disease (EVD) is caused by multiple species of Ebolavirus. Monoclonal antibodies (mAbs) against the virus glycoprotein (GP) are the only class of therapeutic approved for treatment of EVD caused by Zaire ebolavirus (EBOV). Therefore, mAbs targeting multiple Ebolavirus species may represent the next generation of EVD therapeutics. Broadly reactive anti-GP mAbs were produced; among these, mAbs 11886 and 11883 were broadly neutralizing in vitro. A 3.0 [A] cryo-electron microscopy structure of EBOV GP bound to both mAbs shows that 11886 binds a novel epitope bridging the glycan cap (GC), 310 pocket and GP2 N-terminus, whereas 11883 binds the receptor binding region (RBR) and GC. In vitro, 11886 synergized with a range of mAbs with epitope specificities spanning the RBR/GC, including 11883. Notably, 11886 increased the breadth of neutralization by partner mAbs against different Ebolavirus species. These data provide a strategic route to design improved mAb-based next-generation EVD therapeutics.

immunology↗

Structure of a SARS-CoV-2 spike S2 subunit in a pre-fusion, open conformation

The 800 million human infections with SARS-CoV-2 and the likely emergence of new variants and additional coronaviruses necessitate a better understanding of the essential spike glycoprotein and the development of immunogens that foster broader and more durable immunity. The S2 fusion subunit is more conserved in sequence, is essential to function, and would be a desirable immunogen to boost broadly reactive antibodies. It is, however, unstable in structure and in its wild-type form, cannot be expressed alone without irreversible collapse into a six-helix bundle. In addition to the irreversible conformational changes of fusion, biophysical measurements indicate that spike also undergoes a reversible breathing action. However, spike in an open, "breathing" conformation has not yet been visualized at high resolution. Here we describe an S2-only antigen, engineered to remain in its relevant, pre-fusion viral surface conformation in the absence of S1. We also describe a panel of natural human antibodies specific for S2 from vaccinated and convalescent individuals. One of these mAbs, from a convalescent individual, afforded a high-resolution cryo-EM structure of the prefusion S2. The structure reveals a complex captured in an "open" conformation with greater stabilizing intermolecular interactions at the base and a repositioned fusion peptide. Together, this work provides an antigen for advancement of next-generation "booster" immunogens and illuminates the likely breathing adjustments of the coronavirus spike.

microbiology↗