An Engineered Disulfide Bond Reversibly Traps the IgE-Fc_3-4 in a Closed, Nonreceptor Binding Conformation

Journal Article · Fri Aug 02 00:00:00 EDT 2013 · Journal of Biological Chemistry

DOI:https://doi.org/10.1074/jbc.M112.407502· OSTI ID:1087769

Wurzburg, Beth A.; Kim, Beomkyu; Tarchevskaya, Svetlana S.; Eggel, Alexander; Vogel, Monique; Jardetzky, Theodore S. ^[1]

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1. Bern

IgE antibodies interact with the high affinity IgE Fc receptor, FcϵRI, and activate inflammatory pathways associated with the allergic response. The IgE-Fc region, comprising the C-terminal domains of the IgE heavy chain, binds FcϵRI and can adopt different conformations ranging from a closed form incompatible with receptor binding to an open, receptor-bound state. A number of intermediate states are also observed in different IgE-Fc crystal forms. To further explore this apparent IgE-Fc conformational flexibility and to potentially trap a closed, inactive state, we generated a series of disulfide bond mutants. Here we describe the structure and biochemical properties of an IgE-Fc mutant that is trapped in the closed, non-receptor binding state via an engineered disulfide at residue 335 (Cys-335). Reduction of the disulfide at Cys-335 restores the ability of IgE-Fc to bind to its high affinity receptor, FcϵRIα. The structure of the Cys-335 mutant shows that its conformation is within the range of previously observed, closed form IgE-Fc structures and that it retains the hydrophobic pocket found in the hinge region of the closed conformation. Locking the IgE-Fc into the closed state with the Cys-335 mutation does not affect binding of two other IgE-Fc ligands, omalizumab and DARPin E2_79, demonstrating selective blocking of the high affinity receptor binding.

• Research Organization: Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
• Sponsoring Organization: OTHERNIH
• OSTI ID: 1087769
• Journal Information: Journal of Biological Chemistry, Vol. 287, Issue (43) ; 10, 2012; ISSN 0021-9258
• Publisher: American Society for Biochemistry and Molecular Biology
• Country of Publication: United States
• Language: ENGLISH