Noncovalent antibody catenation on a target surface greatly increases the antigen-binding avidity.

Immunoglobulin G (IgG) antibodies are widely used for diagnosis and therapy. Given the unique dimeric structure of IgG, we hypothesized that, by genetically fusing a homodimeric protein (catenator) to the C-terminus of IgG, reversible catenation of antibody molecules could be induced on a surface where target antigen molecules are abundant, and that it could be an effective way to greatly enhance the antigen-binding avidity. A thermodynamic simulation showed that quite low homodimerization affinity of a catenator, e.g. dissociation constant of 100 µM, can enhance nanomolar antigen-binding avidity to a picomolar level, and that the fold enhancement sharply depends on the density of the antigen. In a proof-of-concept experiment where antigen molecules are immobilized on a biosensor tip, the C-terminal fusion of a pair of weakly homodimerizing proteins to three different antibodies enhanced the antigen-binding avidity by at least 110 or 304 folds from the intrinsic binding avidity. Compared with the mother antibody, Obinutuzumab(Y101L) which targets CD20, the same antibody with fused catenators exhibited significantly enhanced binding to SU-DHL5 cells. Together, the homodimerization-induced antibody catenation would be a new powerful approach to improve antibody applications, including the detection of scarce biomarkers and targeted anticancer therapies.

Corrigendum: Structural basis for the broad and potent cross-reactivity of an N501Y-centric antibody against sarbecoviruses.

[This corrects the article DOI: 10.3389/fimmu.2022.1049867.].

Structural basis for the broad and potent cross-reactivity of an N501Y-centric antibody against sarbecoviruses

More than 80% of SARS-CoV-2 variants, including Alpha and Omicron, contain an N501Y mutation in the receptor-binding domain (RBD) of the spike protein. The N501Y change is an adaptive mutation enabling tighter interaction with the human ACE2 receptor. We have developed a broadly neutralizing antibody (nAb), D27LEY, whose binding affinity was intentionally optimized for Y501. This N501Y-centric antibody not only interacts with the Y501-containing RBDs of SARS-CoV-2 variants, including Omicron, with pico- or subnanomolar binding affinity, but also binds tightly to the RBDs with a different amino acid at residue 501. The crystal structure of the Fab fragment of D27LEY bound to the RBD of the Alpha variant reveals that the Y501-containing loop adopts a ribbon-like topology and serves as a small but major epitope in which Y501 is a part of extensive intermolecular interactions. A hydrophobic cleft on the most conserved surface of the RBD core serves as another major binding epitope. These data explain the broad and potent cross-reactivity of this N501Y-centric antibody, and suggest that a vaccine antigenic component composed of the RBD core and a part of receptor-binding motif (RBM) containing tyrosine at residue 501 might elicit broad and potent humoral responses across sarbecoviruses.

Development of Spike Receptor-Binding Domain Nanoparticles as a Vaccine Candidate against SARS-CoV-2 Infection in Ferrets.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a causative agent of the CoV disease 2019 (COVID-19) pandemic, enters host cells via the interaction of its receptor-binding domain (RBD) of the spike protein with host angiotensin-converting enzyme 2 (ACE2). Therefore, the RBD is a promising vaccine target to induce protective immunity against SARS-CoV-2 infection. In this study, we report the development of an RBD protein-based vaccine candidate against SARS-CoV-2 using self-assembling Helicobacter pylori-bullfrog ferritin nanoparticles as an antigen delivery system. RBD-ferritin protein purified from mammalian cells efficiently assembled into 24-mer nanoparticles. Sixteen- to 20-month-old ferrets were vaccinated with RBD-ferritin nanoparticles (RBD nanoparticles) by intramuscular or intranasal inoculation. All vaccinated ferrets with RBD nanoparticles produced potent neutralizing antibodies against SARS-CoV-2. Strikingly, vaccinated ferrets demonstrated efficient protection from SARS-CoV-2 challenge, showing no fever, body weight loss, or clinical symptoms. Furthermore, vaccinated ferrets showed rapid clearance of infectious virus in nasal washes and lungs as well as of viral RNA in respiratory organs. This study demonstrates that spike RBD-nanoparticles are an effective protein vaccine candidate against SARS-CoV-2.