Abstract
Understanding where antibodies recognize antigens can help define mechanisms of action and provide insights into progression of immune responses. We investigate the extent to which information about binding specificity implicitly encoded in amino acid sequence can be leveraged to identify antibody epitopes. In computationally-driven epitope localization, possible antibody-antigen binding modes are modeled, and targeted panels of antigen variants are designed to experimentally test these hypotheses. Prospective application of this approach to two antibodies enabled epitope localization using five or fewer variants per antibody, or alternatively, a six-variant panel for both simultaneously. Retrospective analysis of a variety of antibodies and antigens demonstrated an almost 90% success rate with an average of three antigen variants, further supporting the observation that the combination of computational modeling and protein design can reveal key determinants of antibody-antigen binding and enable efficient studies of collections of antibodies identified from polyclonal samples or engineered libraries.
Highlights
Antibodies have long been recognized for their beneficial roles in vaccination, infection, and clinical therapy, as well as their pathogenic roles in autoimmunity
As we demonstrate in prospective application to two Abs against a tumor antigen, as well as thorough retrospective testing with a wide range of Ab-Ag pairs, Ab sequence and Ag structure alone are sufficient to drive efficient targeting of experimental effort to effectively localize epitopes
We explored the relationship between allowed inter-mutation distance vs. resolution and success rate, since closer mutations could lead to more precise identification of the epitope but at the expense of hitting epitopes less frequently
Summary
Antibodies have long been recognized for their beneficial roles in vaccination, infection, and clinical therapy, as well as their pathogenic roles in autoimmunity. The protective and/or pathogenic capacity of an antibody (Ab) is functionally delimited by the specific epitope(s) that it recognizes on an antigen (Ag). Even Abs targeting the same Ag have demonstrated variable efficacy dependent upon their epitope specificities. Abs against particular epitopes have been associated with protection in the setting of vaccination (Zolla-Pazner et al, 2014; Gottardo et al, 2013; Steel et al, 2010; Gocnık et al, 2007; Liu et al, 2016; Eggink et al, 2014; Margine et al, 2013) and natural infection (Walker et al, 2010; Lu et al, 2016). The identification of epitopes contributing to potent antibody bioactivity is rapidly gaining attention in vaccine design efforts (Haynes, 2015; West et al, 2014; Zolla-Pazner et al, 2016; Correia et al, 2014; He et al, 2015; Lanzavecchia et al, 2016; Pica and Palese, 2013; Subbarao and Matsuoka, 2013) as well as in reducing immunogenic responses against protein drug candidates (Nagata and Pastan, 2009; Onda et al, 2011; Onda et al, 2008)
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