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The Rad50 zinc-hook is a structure joining Mre11 complexes in DNA recombination and repair

Nature volume 418, pages 562–566 (2002)Cite this article

Abstract

The Mre11 complex (Mre11–Rad50–Nbs1) is central to chromosomal maintenance and functions in homologous recombination, telomere maintenance and sister chromatid association1,2,3,4,5,6,7. These functions all imply that the linked binding of two DNA substrates occurs, although the molecular basis for this process remains unknown. Here we present a 2.2 Å crystal structure of the Rad50 coiled-coil region that reveals an unexpected dimer interface at the apex of the coiled coils in which pairs of conserved Cys-X-X-Cys motifs form interlocking hooks that bind one Zn2+ ion. Biochemical, X-ray and electron microscopy data indicate that these hooks can join oppositely protruding Rad50 coiled-coil domains to form a flexible bridge of up to 1,200 Å. This suggests a function for the long insertion in the Rad50 ABC-ATPase domain8. The Rad50 hook is functional, because mutations in this motif confer radiation sensitivity in yeast and disrupt binding at the distant Mre11 nuclease interface. These data support an architectural role for the Rad50 coiled coils in forming metal-mediated bridging complexes between two DNA-binding heads. The resulting assemblies have appropriate lengths and conformational properties to link sister chromatids in homologous recombination and DNA ends in non-homologous end-joining.

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Figure 1: Rad50 domains, sequence conservation of the CXXC motif and Zn2+-mediated dimerization of Rad50.
Figure 2: Crystal structures of the central portion of the Rad50 coiled coil reveal a dimerization motif mediated by a single Zn2+-binding site.
Figure 3: Negatively stained electron micrographs of human and P. furiosus Mre11 complexes reveal conformational properties of the component domains including Zn2+-mediated tail-to-tail linkages between individual coiled-coil arms of Rad50.
Figure 4: In vivo effects of cysteine mutations in the CXXC motif and biological implications for metal-mediated dimerization of Rad50.

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Acknowledgements

We thank D. Turk for MAIN; M. Gehl for atomic absorption spectroscopy analysis and A.-M. Hayes for technical advice. We acknowledge the key facilities and staff of synchrotron beamlines BL9-2 (SSRL) and 5.0.2 (ALS). Key funding was provided by the US Department of Energy (DOE) and NCI (J.A.T. and J.P.C), the American Cancer Society (J.P.C.), the Canadian Institutes of Health Research (L.C.), National Institutes of Health, Human Frontiers Science Program and DOE (J.H.J.P), Skaggs Institute for Chemical Biology (K.P.H.), the National Institutes of Mental Health (C.T.M.) and The Swiss National Science Foundation (J.L.B).

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Author notes

  1. Karl-Peter Hopfner and Lisa Craig: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Molecular Biology and Skaggs Institute for Chemical Biology, The Scripps Research Institute, La Jolla, California, 92037, USA

    Lisa Craig, Gabriel Moncalian, Jean-Luc Bodmer & John A. Tainer

  2. Gene Center and Institute of Biochemistry, University of Munich, 81377, Munich, Germany

    Karl-Peter Hopfner & Annette Karcher

  3. Laboratory of Genetics, University of Wisconsin, Madison, Wisconsin, 53706, USA

    Robert A. Zinkel

  4. Molecular Biology, Memorial Sloan-Kettering Cancer Center, New York, New York, 10021, USA

    Takehiko Usui & John H. J. Petrini

  5. Department of Molecular Pharmacology and Experimental Therapeutics, Mayo Clinic and Foundation, Rochester, Minnesota, 55905, USA

    Barbara A. L. Owen & Cynthia T. McMurray

  6. The Radiation Oncology Research Laboratory, University of Maryland School of Medicine, Baltimore, Maryland, 21201, USA

    Brendan Henderson & James P. Carney

  7. Life Sciences Division, Lawrence Berkeley National Laboratory, Berkeley, California, 94720, USA

    John A. Tainer

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Correspondence to John H. J. Petrini or John A. Tainer.

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Hopfner, KP., Craig, L., Moncalian, G. et al. The Rad50 zinc-hook is a structure joining Mre11 complexes in DNA recombination and repair. Nature 418, 562–566 (2002). https://doi.org/10.1038/nature00922

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