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Structure of a repair enzyme interrogating undamaged DNA elucidates recognition of damaged DNA

Nature volume 434, pages 612–618 (2005)Cite this article

Abstract

How DNA repair proteins distinguish between the rare sites of damage and the vast expanse of normal DNA is poorly understood. Recognizing the mutagenic lesion 8-oxoguanine (oxoG) represents an especially formidable challenge, because this oxidized nucleobase differs by only two atoms from its normal counterpart, guanine (G). Here we report the use of a covalent trapping strategy to capture a human oxoG repair protein, 8-oxoguanine DNA glycosylase I (hOGG1), in the act of interrogating normal DNA. The X-ray structure of the trapped complex features a target G nucleobase extruded from the DNA helix but denied insertion into the lesion recognition pocket of the enzyme. Free energy difference calculations show that both attractive and repulsive interactions have an important role in the preferential binding of oxoG compared with G to the active site. The structure reveals a remarkably effective gate-keeping strategy for lesion discrimination and suggests a mechanism for oxoG insertion into the hOGG1 active site.

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Figure 1: Generation of 8-oxoguanine (oxoG), its recognition by human 8-oxoguanine DNA glycosylase (hOGG1) and overview of the structure-based trapping strategy used here to obtain a complex of hOGG1 bound to undamaged DNA.
Figure 2: Comparison of the overall structures of trapped complexes obtained with oxoG-containing (left) or G-containing (right) DNA.
Figure 3: View of the active site region of hOGG1–DNA complexes, showing extra-helical nucleobases bound to either the lesion recognition pocket or the alternative site.
Figure 4: Computational analysis of the binding free energy difference between oxoG and G.
Figure 5: Superposition of the oxoG complex with the G complex in the region around the protein–DNA interface.

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Acknowledgements

We are grateful to Y. Korkhin for help in data collection and processing. We thank Enanta Pharmaceuticals for use of their X-ray instrumentation. We acknowledge the entire staff at MACCHESS, especially C. Heaton and B. Miller, and NSLS X4A for assistance in data collection and processing. We thank D. Jeruzalmi and C. Fromme for valuable discussions. This work was supported by grants from the NIH to G.L.V. and M.K.Author contributions A.B. was responsible for performing the structural and biochemical experiments described herein, whereas W.Y. performed the computational simulations.

Author information

Authors and Affiliations

  1. Departments of Chemistry and Chemical Biology,

    Anirban Banerjee, Wei Yang, Martin Karplus & Gregory L. Verdine

  2. Molecular and Cellular Biology, Harvard University, 12 Oxford Street, Cambridge, Massachusetts, 02138, USA

    Gregory L. Verdine

  3. L'Institut de Science et d'Ingénierie Supramoléculaires (ISIS), Université Louis Pasteur, 8 allée Gaspard Monge, 67000, Strasbourg, France

    Martin Karplus

Authors
  1. Anirban Banerjee
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Corresponding authors

Correspondence to Martin Karplus or Gregory L. Verdine.

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The authors declare that they have no competing financial interests.

Supplementary information

Supplementary Table S1

Data collection and model statistics. (DOC 54 kb)

Supplementary Figure Legends

Figure captions for Supplementary Figures S1-S4. (DOC 35 kb)

Supplementary Methods

Additional details on computational methods and data collection and structure solution. (DOC 56 kb)

Supplementary Figure S1

Structural validation of the trapping strategy and crosslinking biochemistry. (PDF 658 kb)

Supplementary Figure S2

Electron density map around the G in the exo- site and the crosslinked C in the G-complex. (JPG 550 kb)

Supplementary Figure S3

Structural representations of the carbonyl of Gly42 and different nucleobases used in this study. (JPG 149 kb)

Supplementary Figure S4

Electron density maps around the analogs in the crosslinked complexes. (JPG 767 kb)

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Banerjee, A., Yang, W., Karplus, M. et al. Structure of a repair enzyme interrogating undamaged DNA elucidates recognition of damaged DNA. Nature 434, 612–618 (2005). https://doi.org/10.1038/nature03458

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