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Structural basis of sequence-specific Holliday junction cleavage by MOC1

Nature Chemical Biology volume 15, pages 1241–1248 (2019)Cite this article

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Abstract

The Holliday junction (HJ) is a key intermediate during homologous recombination and DNA double-strand break repair. Timely HJ resolution by resolvases is critical for maintaining genome stability. The mechanisms underlying sequence-specific substrate recognition and cleavage by resolvases remain elusive. The monokaryotic chloroplast 1 protein (MOC1) specifically cleaves four-way DNA junctions in a sequence-specific manner. Here, we report the crystal structures of MOC1 from Zea mays, alone or bound to HJ DNA. MOC1 uses a unique β-hairpin to embrace the DNA junction. A base-recognition motif specifically interacts with the junction center, inducing base flipping and pseudobase-pair formation at the strand-exchanging points. Structures of MOC1 bound to HJ and different metal ions support a two-metal ion catalysis mechanism. Further molecular dynamics simulations and biochemical analyses reveal a communication between specific substrate recognition and metal ion-dependent catalysis. Our study thus provides a mechanism for how a resolvase turns substrate specificity into catalytic efficiency.

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Fig. 1: MOC1 cleaves HJ DNA in a sequence-specific manner.
Fig. 2: Structures of ZmMOC1 and its complex with a HJ.
Fig. 3: Interactions between MOC1 and HJ DNA backbones.
Fig. 4: Base-specific recognition.
Fig. 5: The active sites of MOC1 and metal ion coordination.
Fig. 6: The communication between substrate binding and metal ion coordination, as revealed by molecular dynamics simulations.

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Data availability

Atomic coordinates and structure factors of apo-MOC1, MOC1–HJ–Ca2+, MOC1D115N–HJ–Mg2+ and MOC1H253A–HJ–Mg2+ have been deposited in the Protein Data Bank (PDB) with accession codes 6IS9, 6JRF, 6IS8 and 6JRG, respectively. All data generated and analyzed during this study are included in this published article. Constructs encoding full-length and truncated ZmMOC1 variants are available from the corresponding author upon reasonable request.

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Acknowledgements

We thank the staff of BL17B, BL18U1 and BL19U1 beamlines at the National Facility for Protein Science Shanghai (NFPS) and Shanghai Synchrotron Radiation Facility, Shanghai, People’s Republic of China, for assistance with X-ray data collection. We thank H. Yu (Howard Hughes Medical Institute, University of Texas Southwestern Medical Center, Dallas, TX) for critical comments on this manuscript and Y. Huang (Fuzhou University, Fujian, China) for assistance with the molecular dynamics simulations. This work is supported by the National Natural Science Foundation of China 31971222 (Z.L.), 21603033 (J.L.), 31370737 (M.H.) and 31670739 (M.H.), and the National Key R&D Program of China 2017YFE0103200 (M.H.).

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  1. These authors contributed equally: Huajian Lin, Danping Zhang, Ke Zuo.

Authors and Affiliations

  1. College of Chemistry, Fuzhou University, Fuzhou, China

    Huajian Lin, Danping Zhang, Ke Zuo, Jinyu Li, Mingdong Huang & Zhonghui Lin

  2. College of Biological Science and Engineering, Fuzhou University, Fuzhou, China

    Cai Yuan

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Contributions

H.L. carried out all the cloning, DNA binding and cleavage experiments, and the protein purification and crystallization of both native and SeMet MOC1. D.Z. carried out the purification and crystallization of MOC1–HJ complexes. H.L. and D.Z. performed the X-ray data collection. C.Y. performed structure refinements. K.Z. carried out the molecular dynamics simulations under the supervision of J.L. M.H. supervised the project and analyzed the data. Z.L. conceived and designed the project, determined the crystal structures and wrote the manuscript with the input of all authors. All authors read and approved the final manuscript.

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Correspondence to Jinyu Li, Mingdong Huang or Zhonghui Lin.

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Lin, H., Zhang, D., Zuo, K. et al. Structural basis of sequence-specific Holliday junction cleavage by MOC1. Nat Chem Biol 15, 1241–1248 (2019). https://doi.org/10.1038/s41589-019-0377-4

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