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An autoinhibited state of 53BP1 revealed by small molecule antagonists and protein engineering
Gaofeng Cui1, Maria Victoria Botuyan1, Pascal Drané2
1Department of Biochemistry and Molecular Biology, Mayo Clinic, Rochester, MN, USA.
Abstract:
The recruitment of 53BP1 to chromatin, mediated by its recognition of histone H4 dimethylated at lysine 20 (H4K20me2), is important for DNA double-strand break repair. Using a series of small molecule antagonists, we demonstrate a conformational equilibrium between an open and a pre-existing lowly populated closed state of 53BP1 in which the H4K20me2 binding surface is buried at the interface between two interacting 53BP1 molecules. In cells, these antagonists inhibit the chromatin recruitment of wild type 53BP1, but do not affect 53BP1 variants unable to access the closed conformation despite preservation of the H4K20me2 binding site. Thus, this inhibition operates by shifting the conformational equilibrium toward the closed state. Our work therefore identifies an auto-associated form of 53BP1-autoinhibited for chromatin binding-that can be stabilized by small molecule ligands encapsulated between two 53BP1 protomers. Such ligands are valuable research tools to study the function of 53BP1 and have the potential to facilitate the development of new drugs for cancer therapy.
Insights
Small molecules stabilize a closed, auto-inhibited state of 53BP1, blocking its DNA repair function. This discovery offers new tools for studying 53BP1 and developing cancer therapies.
Area of Science:
- Molecular Biology
- Biochemistry
- Cancer Research
Background:
- 53BP1 protein is crucial for DNA double-strand break repair.
- Its recruitment to chromatin depends on recognizing histone H4K20me2.
- Understanding 53BP1 regulation is key for cancer therapy development.
Purpose of the Study:
- To investigate the conformational states of 53BP1.
- To identify mechanisms regulating 53BP1 chromatin binding.
- To explore small molecule modulators of 53BP1 function.
Main Methods:
- Utilized small molecule antagonists to probe 53BP1 conformation.
- Assessed chromatin recruitment of wild-type and variant 53BP1 in cells.
- Investigated protein-protein interactions and conformational equilibrium.
Main Results:
- Identified a conformational equilibrium between open and closed states of 53BP1.
- Demonstrated that small molecules stabilize a closed, auto-inhibited 53BP1 conformation.
- Showed inhibition of chromatin recruitment by stabilizing the closed state.
Conclusions:
- 53BP1 exists in an auto-inhibited closed state, inaccessible for chromatin binding.
- Small molecule ligands can stabilize this closed state, inhibiting DNA repair.
- These findings provide novel research tools and potential therapeutic strategies for cancer.
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