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Characterizing the Structure and Dynamics of Adaptive Immune Proteins to Inform Immunotherapy Design Helen LeClair Woodward
- Format:
- Book
- Thesis/Dissertation
- Author/Creator:
- Woodward, Helen LeClair, author.
- Language:
- English
- Subjects (All):
- Biophysics.
- Immunology.
- Biochemistry.
- Cellular biology.
- 0786.
- 0982.
- 0487.
- 0379.
- Local Subjects:
- Biophysics.
- Immunology.
- Biochemistry.
- Cellular biology.
- 0786.
- 0982.
- 0487.
- 0379.
- Physical Description:
- 1 electronic resource (171 pages)
- Contained In:
- Dissertations Abstracts International 86-12B
- Place of Publication:
- Ann Arbor : ProQuest Dissertations and Theses, 2025
- Language Note:
- English
- Summary:
- Human interleukin-2 (IL-2) is a crucial cytokine for T cell regulation, with therapeutic potential in cancer and autoimmune diseases. However, IL-2's pleiotropic effects across different immune cell types often lead to toxicity and limited efficacy. The first study in this thesis presents a detailed characterization of IL-2 dynamics compared to two engineered IL-2 mutants, "superkines" S15 and S1, which exhibit biased signaling towards effector T cells. Using NMR spectroscopy and molecular dynamics simulations, we demonstrate significant variations in core dynamic landscapes across the three variants. Furthermore, we rationally design a mutation (L56A) in the S1 core network, which partially reverts its dynamics, receptor binding affinity, and T cell signaling behavior. Our results suggest that modulating IL-2 dynamics is an untapped approach for designing immunotherapies with improved immune cell selectivity profiles.The intrinsic instability and inconsistent T cell receptor binding mode of the class I major histocompatibility complex (MHC-I) are fundamental challenges that hinder development of therapeutics. The second study leverages the positive allosteric coupling between the peptide and β2m subunits of MHC-I by engineering a disulfide bond bridging conserved epitopes across the HC/β2m interface to generate stable, peptide-receptive molecules. Using solution NMR, we demonstrate that the disulfide bond induces long-range conformational and dynamical changes in the α2-1 helix., indicating that the interchain disulfide bond stabilizes MHC-I molecules in an open conformation to promote peptide exchange. In the third study, we demonstrate the application of a designed MHC-I design, SMART A*02:01, for solution mapping of the A6c134 TCR docking footprint using NMR spectroscopy. We establish that the use of our engineered construct recapitulates the native conformation of the MHC-I peptide binding groove and accurately detects the critical binding sites on SMART A*02:01 that engage A6c134. Our approach allows for high-resolution mapping of TCR/MHC-I interactions, alleviating the need for complete structure elucidation, and may be paired with computational modeling approaches for structure-guided optimization and clinical development.Taken together, solution NMR techniques provide high-resolution insights into protein structure and conformational dynamics. Furthermore, these efforts highlight the application of NMR analyses to inform rational protein design and guide the development of diagnostics and therapeutics
- Notes:
- Source: Dissertations Abstracts International, Volume: 86-12, Section: B.
- Advisors: Sgourakis, Nikolaos G. Committee members: Bowman, Gregory R.; Lynch, Kristen W.; Rhoades, Elizabeth; Spangler, Jamie B.; Sharp, Kim A.
- Ph.D. University of Pennsylvania 2025
- Local Notes:
- School code: 0175
- ISBN:
- 9798280760233
- Access Restriction:
- Restricted for use by site license
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