Abstract
Holliday proposed a four-way DNA junction as an intermediate in homologous recombination1, and such Holliday junctions have since been identified as a central component in DNA recombination and repair2. Phage T4 endonuclease VII (endo VII) was the first enzyme shown to resolve Holliday junctions into duplex DNAs by introducing symmetrical nicks in equivalent strands3. Several Holliday junction resolvases have since been characterized4, but an atomic structure of a resolvase complex with a Holliday junction remained elusive. Here we report the crystal structure of an inactive T4 endo VII(N62D) complexed with an immobile four-way junction with alternating arm lengths of 10 and 14 base pairs. The junction is a hybrid of the conventional square-planar and stacked-X conformation. Endo VII protrudes into the junction point from the minor groove side, opening it to a 14 Å × 32 Å parallelogram. This interaction interrupts the coaxial stacking, yet every base pair surrounding the junction remains intact. Additional interactions involve the positively charged protein and DNA phosphate backbones. Each scissile phosphate that is two base pairs from the crossover interacts with a Mg2+ ion in the active site. The similar overall shape and surface charge potential of the Holliday junction resolvases endo VII, RuvC, Ydc2, Hjc and RecU, despite having different folds, active site composition and DNA sequence preference, suggest a conserved binding mode for Holliday junctions.
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Acknowledgements
We thank D. Leahy and M. Gellert for critical reading of the manuscript, S. Ramon-Maiques for collecting the diffracting data of the endo VII–heteroduplex complex, and D. M. Lilley for background reading materials. C.B. thanks J. Basquin, E. Ennifar and C. Sauter for help with crystallization and data collection, and M. Nowotny and J. Y. Lee for help with manuscript preparation. This research was supported by EMBL, Deutsche Forschungsgemeinschaft and the Intramural Research Program of NIDDK, NIH.
Author Contributions C.B. carried out all experiments. The project was initiated at EMBL and finished at NIH. All authors contributed to experimental design, interpretation and manuscript preparation.
Atomic coordinates and structure factors of the endo VII–DNA complexes have been deposited in the Protein Data Bank. The accession codes are 2QNC and 2QNF for the Holliday junction and the heteroduplex complex, respectively.
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Biertümpfel, C., Yang, W. & Suck, D. Crystal structure of T4 endonuclease VII resolving a Holliday junction. Nature 449, 616–620 (2007). https://doi.org/10.1038/nature06152
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DOI: https://doi.org/10.1038/nature06152
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