Protein-Protein Recognition: The Neonatal Fc Receptor and Immunoglobulin G

Author: Martin, Warham Lance

Year: 2001

Degree: Dissertation (Ph.D.)

Advisors: Bjorkman, Pamela J.; Meyerowitz, Elliot M.

Committee Members: Rees, Douglas C.; Bjorkman, Pamela J.; Fraser, Scott E.; Rothenberg, Ellen V.; Meyerowitz, Elliot M.

Option: Biochemistry and Molecular Biophysics

DOI: 10.7907/9ek6-6833

Abstract

The neonatal Fc receptor (FcRn) binds the Fc portion of immunoglobulin G (IgG) at the acidic pH of endosomes or the gut and releases IgG at the alkaline pH of blood. FcRn is responsible for the maternofetal transfer of IgG and for rescuing endocytosed IgG from a default degradative pathway. We investigated how FcRn interacts with IgG by constructing a heterodimeric form of the Fc (hdFc) that contains one FcRn binding site. This molecule was used to characterize the interaction between one FcRn molecule and one Fc and to determine under what conditions FcRn forms a dimer. The hdFc binds one FcRn molecule at pH 6.0 with a Kd of 80 nM. In solution and with FcRn anchored to solid supports, the heterodimeric Fc does not induce a dimer of FcRn molecules. FcRnhdFc complex crystals were obtained and the complex structure was solved to 2.8 Å resolution. Analysis of this structure refined the understanding of the mechanism of the pH-dependent binding, shed light on the role played by carbohydrates in the Fc binding, and provided insights on how to design therapeutic IgG antibodies with longer serum half-lives. The FcRn-hdFc complex in the crystal did not contain the FcRn dimer. To characterize the tendency of FcRn to form a dimer in a membrane we analyzed the tendency of the hdFc to induce cross-phosphorylation of FcRn-tyrosine kinase chimeras. We also constructed FcRn-cyan and FcRn-yellow fluorescent proteins and have analyzed the tendency of these molecules to exhibit fluorescence resonance energy transfer. As of now, neither of these analyses have lead to conclusive results. In the process of acquiring the context to appreciate the structure of the FcRn-hdFc interface, we developed a study of 171 other nonobligate protein-protein interfaces that includes an original principal component analysis of the quantifiable aspects of these interfaces.

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