IgE antibody dynamics and its role in receptor binding and inhibition
Abstract/Contents
- Abstract
- IgE antibodies interact with the high affinity IgE Fc receptor (Fc [epsilon] RI) expressed on mast cells and basophils, and the low affinity IgE Fc receptor (CD23) on B cells. These interactions are directly involved in triggering the allergen-specific activation of inflammatory responses. The IgE-Fc region, comprising the C-terminal domains of the IgE heavy chain, binds Fc [epsilon] RI and the crystal structures of the human IgE-Fc alone and IgE-Fc complexes exhibited IgE-Fc conformational flexibility. In addition, the crystal structure of IgE-Fc bound to CD23 indicated a preference of CD23 for a closed conformation of the IgE-Fc incompatible with Fc [epsilon] RI binding. These observations suggested the importance of IgE conformational dynamics for its receptor binding mechanism and the potential to identify non-classical or allosteric inhibitors for IgE-receptor interactions. To investigate IgE dynamics and its role in receptor binding and inhibition, I have employed 5 experimental approaches: 1) IgE-Fc mutants trapped by an engineered disulfide bond, blocking Fc [epsilon] RI but not IgE inhibitor binding, 2) a fluorescence assay for IgE-Fc:Fc [epsilon] RI binding and inhibition that can distinguish IgE-inhibitors based on their proximity to the Fc [epsilon] RI binding site, 3) the determination of the E2_79 crystal structure showing that the IgE inhibitor E2_79 binds IgE-Fc outside the Fc [epsilon] RI-binding site and acts through a facilitated dissociation mechanism to block receptor binding, 4) the development of a TR-FRET assay suitable for high-throughput screening for small molecule inhibitors of IgE-receptor binding, and 5) a single-molecule FRET approach to characterize the conformational states of the IgE-Fc3-4 and observe changes upon Fc [epsilon] RI, CD23, and E2_79. These experiments revealed different conformational requirements for IgE binding, identified a disruptive inhibitor that actively dissociates preformed receptor complexes, and provided direct observation of receptor binding-induced IgE-Fc conformational changes. This work provides a new perspective on the role of IgE dynamics in receptor binding and inhibition, and makes possible the development of new disruptive inhibitors for IgE-receptor binding acting through non-classical or allosteric mechanisms.
Description
| Type of resource | text |
|---|---|
| Form | electronic; electronic resource; remote |
| Extent | 1 online resource. |
| Publication date | 2013 |
| Issuance | monographic |
| Language | English |
Creators/Contributors
| Associated with | Kim, Beomkyu |
|---|---|
| Associated with | Stanford University, Department of Structural Biology. |
| Primary advisor | Jardetzky, Theodore |
| Thesis advisor | Jardetzky, Theodore |
| Thesis advisor | Jones, Patricia P |
| Thesis advisor | Puglisi, Joseph D |
| Thesis advisor | Weis, William I |
| Advisor | Jones, Patricia P |
| Advisor | Puglisi, Joseph D |
| Advisor | Weis, William I |
Subjects
| Genre | Theses |
|---|
Bibliographic information
| Statement of responsibility | Beomkyu Kim. |
|---|---|
| Note | Submitted to the Department of Structural Biology. |
| Thesis | Ph.D. Stanford University 2013 |
| Location | electronic resource |
Access conditions
- Copyright
- © 2013 by Beomkyu Kim
- License
- This work is licensed under a Creative Commons Attribution Non Commercial 3.0 Unported license (CC BY-NC).
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