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Engineering the Fab fragment of the anti-IgE omalizumab to prevent Fab crystallization and permit IgE-Fc complex crystallization

Research output: Contribution to journalArticle

Original languageEnglish
JournalActa Crystallographica Section F:Structural Biology Communications
Publication statusPublished - 3 Feb 2020


King's Authors


Immunoglobulin E (IgE) plays a central role in the allergic response, in which
cross-linking of the allergen by FcεRI-bound IgE triggers mast-cell and basophil
degranulation and the release of inflammatory mediators. The high-affinity
interaction between IgE and FcεRI is a long-standing target for therapeutic
intervention in allergic disease. Omalizumab is a clinically approved anti-IgE
monoclonal antibody that binds to free IgE, also with high affinity, preventing its
interaction with FcεRI. All attempts to crystallize the pre-formed complex
between the omalizumab Fab and the Fc region of IgE (IgE-Fc), in order to
understand the structural basis for its mechanism of action, surprisingly failed.
Instead, the Fab alone selectively crystallized in different crystal forms, and their
structures revealed intermolecular Fab–Fab interactions that were clearly strong
enough to disrupt the Fab–IgE-Fc complexes. Some of these interactions were
common to other Fab crystal structures. Mutations were therefore designed to
disrupt two recurring packing interactions observed in the omalizumab Fab
crystal structures without interfering with the ability of the omalizumab Fab to
recognize IgE-Fc; this led to the successful crystallization and subsequent
structure determination of the Fab–IgE-Fc complex. The mutagenesis strategy
adopted to achieve this result is applicable to other intractable Fab–antigen
complexes or systems in which Fabs are used as crystallization chaperones.

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