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Sequence selectivity and degeneracy of a restriction endonuclease mediated by DNA intercalation

Nature Structural Biology volume 9, pages 42–47 (2002)Cite this article

Abstract

The crystal structure of the HincII restriction endonuclease–DNA complex shows that degenerate specificity for blunt-ended cleavage at GTPyPuAC sequences arises from indirect readout of conformational preferences at the center pyrimidine-purine step. Protein-induced distortion of the DNA is accomplished by intercalation of glutamine side chains into the major groove on either side of the recognition site, generating bending by either tilt or roll at three distinct loci. The intercalated side chains propagate a concerted shift of all six target-site base pairs toward the minor groove, producing an unusual cross-strand purine stacking at the center pyrimidine–purine step. Comparison of the HincII and EcoRV cocrystal structures suggests that sequence-dependent differences in base–stacking free energies are a crucial underlying factor mediating protein recognition by indirect readout.

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Figure 1: Crystal structure of the HincII–DNA complex.
Figure 2: Enzyme–DNA interactions.
Figure 3: DNA conformation.

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Acknowledgements

This work was supported by the NIH (to J.J.P.), and by an American Cancer Society Postdoctoral Fellowship and a NSF-POWRE award (to N.C.H.). Portions of this research were carried out at the Stanford Synchrotron Radiation Laboratory, a national user facility operated by Stanford University on behalf of the U.S. Department of Energy, Office of Basic Energy Sciences. The SSRL Structural Molecular Biology Program is supported by the Department of Energy, Office of Biological and Environmental Research, and by the National Institutes of Health, National Center for Research Resouces, Biomedical Technology Program, and the National Institute of General Medical Sciences.

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  1. Nancy C. Horton

    Present address: Department of Biochemistry and Molecular Biophysics, College of Medicine, University of Arizona, Tucson, Arizona, 85724, USA

Authors and Affiliations

  1. Department of Chemistry and Biochemistry, and Interdepartmental Program in Biomolecular Science and Engineering, University of California at Santa Barbara, Santa Barbara, 93106-9510, California, USA

    Nancy C. Horton & John J. Perona

  2. New England Biolabs, 32 Tozer Road, Beverly, 01915, Massachusetts, USA

    Lydia F. Dorner

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Correspondence to John J. Perona.

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Horton, N., Dorner, L. & Perona, J. Sequence selectivity and degeneracy of a restriction endonuclease mediated by DNA intercalation. Nat Struct Mol Biol 9, 42–47 (2002). https://doi.org/10.1038/nsb741

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