6ZWK
Crystal structure of the phosphorylated C-terminal tail of histone H2AX in complex with a specific nanobody (C6 gammaXbody)
Summary for 6ZWK
| Entry DOI | 10.2210/pdb6zwk/pdb |
| Descriptor | gammaXbody, Histone H2AX, SODIUM ION, ... (5 entities in total) |
| Functional Keywords | nanobody, phosphopeptide-recognition protein, gamma-h2ax, immune system |
| Biological source | Vicugna pacos (alpaca) More |
| Total number of polymer chains | 12 |
| Total formula weight | 106914.98 |
| Authors | McEwen, A.G.,Moeglin, E.,Desplancq, D.,Weiss, E.,Poterszman, A. (deposition date: 2020-07-28, release date: 2021-07-21, Last modification date: 2024-10-23) |
| Primary citation | Moeglin, E.,Desplancq, D.,Stoessel, A.,Massute, C.,Ranniger, J.,McEwen, A.G.,Zeder-Lutz, G.,Oulad-Abdelghani, M.,Chiper, M.,Lafaye, P.,Di Ventura, B.,Didier, P.,Poterszman, A.,Weiss, E. A Novel Nanobody Precisely Visualizes Phosphorylated Histone H2AX in Living Cancer Cells under Drug-Induced Replication Stress. Cancers (Basel), 13:-, 2021 Cited by PubMed Abstract: Histone H2AX phosphorylated at serine 139 (γ-H2AX) is a hallmark of DNA damage, signaling the presence of DNA double-strand breaks and global replication stress in mammalian cells. While γ-H2AX can be visualized with antibodies in fixed cells, its detection in living cells was so far not possible. Here, we used immune libraries and phage display to isolate nanobodies that specifically bind to γ-H2AX. We solved the crystal structure of the most soluble nanobody in complex with the phosphopeptide corresponding to the C-terminus of γ-H2AX and show the atomic constituents behind its specificity. We engineered a bivalent version of this nanobody and show that bivalency is essential to quantitatively visualize γ-H2AX in fixed drug-treated cells. After labelling with a chemical fluorophore, we were able to detect γ-H2AX in a single-step assay with the same sensitivity as with validated antibodies. Moreover, we produced fluorescent nanobody-dTomato fusion proteins and applied a transduction strategy to visualize with precision γ-H2AX foci present in intact living cells following drug treatment. Together, this novel tool allows performing fast screenings of genotoxic drugs and enables to study the dynamics of this particular chromatin modification in individual cancer cells under a variety of conditions. PubMed: 34282773DOI: 10.3390/cancers13133317 PDB entries with the same primary citation |
| Experimental method | X-RAY DIFFRACTION (1.55 Å) |
Structure validation
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