Redesigning an FKBP–ligand interface to generate chemical dimerizers with novel specificity
Open accessAbstract
FKBP ligand homodimers can be used to activate signaling events inside cells and animals that have been engineered to express fusions between appropriate signaling domains and FKBP. However, use of these dimerizers in vivo is potentially limited by ligand binding to endogenous FKBP. We have designed ligands that bind specifically to a mutated FKBP over the wild-type protein by remodeling an FKBP-ligand interface to introduce a specificity binding pocket. A compound bearing an ethyl substituent in place of a carbonyl group exhibited sub-nanomolar affinity and 1,000-fold selectivity for a mutant FKBP with a compensating truncation of a phenylalanine residue. Structural and functional analysis of the new pocket showed that recognition is surprisingly relaxed, with the modified ligand only partially filling the engineered cavity. We incorporated the specificity pocket into a fusion protein containing FKBP and the intracellular domain of the Fas receptor. Cells expressing this modified chimeric protein potently underwent apoptosis in response to AP1903, a homodimer of the modified ligand, both in culture and when implanted into mice. Remodeled dimerizers such as AP1903 are ideal reagents for controlling the activities of cells that have been modified by gene therapy procedures, without interference from endogenous FKBP.
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Koduri and colleagues demonstrated inducible degradation of multiple fusion proteins linked to a 25-amino acid zinc finger degron tag derived from IKZF3 and a >50% reduction in luminescence from tumour cells expressing a degron-tagged luciferase transgene upon treatment with thalidomide analogues.
“Notably, Koduri and colleagues205 have demonstrated inducible degradation of multiple fusion proteins linked to a 25-amino acid zinc finger degron tag derived from IKZF3 and a >50% reduction in luminescence from tumour cells expressing a degron-tagged luciferase transgene upon treatment with thalidomide analogues.”
From Cancer therapies based on targeted protein degradation — lessons learned with lenalidomide (2021) · cites this paper as [205]
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- Authors
- Clackson, T., Yang, W., Rozamus, L. W., Hatada, M., Amara, J. F., Rollins, C. T., Stevenson, L. F., Magari, S. R., Wood, S. A., Courage, N. L., Lu, X., Cerasoli, F., Gilman, M., & Holt, D. A.
- Affiliations
- ARIAD Gene Therapeutics, Inc., 26 Landsdowne Street, Cambridge, MA 02139
- ORCID
- Stevenson, Lauren F.
- Venue
- Proceedings of the National Academy of Sciences
- Publisher
- National Academy of Sciences
- Published
- 1998-09
- Volume
- 95
- Issue
- 18
- Pages
- 10437-10442
- ISSN
- 0027-8424 (ISSN-L)
- 1091-6490
- PMC
- PMC27912
- OpenAlex
- W2135450099
- Cited by
- 558
- References
- 33
- Citations by year
- 2026: 28 · 2025: 29 · 2024: 28 · 2023: 22 · 2022: 33 · 2021: 29 · 2020: 26 · 2019: 19 · 2018: 20 · 2017: 21 · 2016: 16 · 2015: 28 · 2014: 22 · 2013: 9 · 2012: 25
- FWCI
- 3.72
- Type
- journal article
- Language
- en
- Open access
- green
- europepmc.org
- Locations
- Proceedings of the National Academy of Sciences (publishedVersion)
- Europe PMC (PubMed Central) (publishedVersion)
- PubMed Central (publishedVersion)
- PubMed (publishedVersion)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Chemical Synthesis and Analysis
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Signaling Pathways in Disease
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Biochemical and Structural Characterization
- Keywords
- FKBP, ligand specificity, specificity pocket, fusion protein, apoptosis, gene therapy
- MeSH terms
- Amino Acid Sequence, Animals, Carrier Proteins, Cell Line, DNA-Binding Proteins, Heat-Shock Proteins, Ligands, Male, Membrane Glycoproteins, Molecular Sequence Data, Molecular Structure, Signal Transduction, Crystallography, X-Ray, Dimerization, Tacrolimus Binding Proteins, Mice, Fas Ligand Protein
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