Convergent Evolution of Neutralizing Antibodies to Staphylococcus aureus γ-Hemolysin C That Recognize an Immunodominant Primary Sequence-Dependent B-Cell Epitope

Abstract

Staphylococcus aureus infection is a major public health threat in part due to the spread of antibiotic resistance and repeated failures to develop a protective vaccine. Infection is associated with production of virulence factors that include exotoxins that attack host barriers and cellular defenses, such as the leukocidin (Luk) family of bicomponent pore-forming toxins. To investigate the structural basis of antibody-mediated functional inactivation of Luk toxins, we generated a panel of murine monoclonal antibodies (MAbs) that neutralize host cell killing by the γ-hemolysin HlgCB. By biopanning these MAbs against a phage-display library of random Luk peptide fragments, we identified a small subregion within the rim domain of HlgC as the epitope for all the MAbs. Within the native holotoxin, this subregion folds into a conserved β-hairpin structure, with exposed key residues, His252 and Tyr253, required for antibody binding. On the basis of the phage-display results and molecular modeling, a 15-amino-acid synthetic peptide representing the minimal epitope on HlgC (HlgC241-255) was designed, and preincubation with this peptide blocked antibody-mediated HIgCB neutralization. Immunization of mice with HlgC241-255 or the homologous LukS246-260 subregion peptide elicited serum antibodies that specifically recognized the native holotoxin subunits. Furthermore, serum IgG from patients who were convalescent for invasive S. aureus infection showed neutralization of HlgCB toxin activity ex vivo, which recognized the immunodominant HlgC241-255 peptide and was dependent on His252 and Tyr253 residues. We have thus validated an efficient, rapid, and scalable experimental workflow for identification of immunodominant and immunogenic leukotoxin-neutralizing B-cell epitopes that can be exploited for new S. aureus-protective vaccines and immunotherapies.

Diversity of Functionally Distinct Clonal Sets of Human Conventional Memory B Cells That Bind Staphylococcal Protein A

“…Whereas earlier reports have been limited to murine studies ( 18 , 27 , 60 ), the current work is the first to document the antigenicity of alternative determinants on SpA for the human immune system. We have recently reported the molecular characterization of immunogenic sites in staphylococcal Leukocidins, which induce in vivo responses that can contribute to functional inactivation of these important toxins ( 69 , 70 ). Akin to our advocacy of these minimal epitopes on Leukocidins for inclusion in multi-module vaccines, our current studies herald a practical approach towards further targeted augmentation of immunity via vaccination by inclusion of alternative immunogenic SpA sites.…”

The power of phages: revolutionizing cancer treatment

“…It has been widely utilized in the development of vaccines against various microorganisms, including cancer causing pathogens ( 112 ). Researchers have employed phage display libraries to identify immunodominant and immunogenic B-cell epitopes that neutralize toxins from pathogens such as Staphylococcus aureus ( 117 ), Escherichia coli ( 112 ), Salmonella typhi ( 118 ), Borrelia burgdorferi ( 119 ), and viruses like SARS-CoV-2 ( 120 ), Ebola Virus ( 121 ), and Zika virus ( 122 ). Engineered filamentous phages have been employed as direct immunogens in various scenarios, such as immunization against Plasmodium falciparum , the parasite responsible for malaria.…”

Emerging Strategies to Combat β-Lactamase Producing ESKAPE Pathogens

“…Using serum from mice immunized with this protein, intense bactericidal activity was observed against A. baumannii strains [ 330 ]. Vaccines built on recombinant proteins have also been developed against S. aureus using extracellular bacterial vesicles coating mesoporous silica nanoparticles [ 329 , 331 , 332 ]. In K. pneumoniae , the in vivo studies in mice model infection and non-human primate model of severe lower respiratory tract infection revealed the unique immunogenic properties of T cell-specific epitopes [ 333 ], recombinant protein vaccine [ 334 ], and polysaccharide capsule type 2 vaccine [ 335 ].…”