1. Skaggs Institute for Chemical Biology and Department of Molecular Biology, The Scripps Research Institute, La Jolla, California 92037, USA
2. Cancer Research UK Carcinogenesis Group, Paterson Institute for Cancer Research, University of Manchester, Manchester, M20 4BX, UK
3. Department of Cellular and Molecular Physiology, Milton S. Hershey Medical Center, Pennsylvania State University College of Medicine, Hershey, Pennsylvania 17033, USA
4. Center for Structural Biology, Department of Molecular and Cellular Biochemistry, University of Kentucky, Lexington, Kentucky 40536, USA
5. NWCRF Institute, Bangor University, Gwynedd LL57 2UW, UK
6. Institute of Human Genetics, Newcastle University, Newcastle-upon-Tyne, NE1 3BZ, UK
7. Centre for Chemical Biology, Department of Chemistry, University of Sheffield, Sheffield S3 7HF, UK
8. Division of Environmental Health Sciences and the Masonic Cancer Center, University of Minnesota, Minneapolis, Minnesota 55455, USA
9. Life Sciences Division, Bioenergy and Structural Biology, Lawrence Berkeley National Laboratory, Berkeley, California 94720, USA
10. Present addresses: Cancer Research UK DNA Damage Response Group, Paterson Institute for Cancer Research, University of Manchester, Manchester, UK (R.K.); Department of Biology, University of Copenhagen, Ole Maaløes Vej 5, DK-2200 Copenhagen N, Denmark (O.F.); Department of Toxicology, University of Mainz, D-55131 Mainz, Germany (B.V.).

Correspondence to: Anthony E. Pegg³John A. Tainer^1,9 Correspondence and requests for materials should be addressed to J.A.T. (Email: jat@scripps.edu) or A.E.P. (Email: aep1@psu.edu).

Abstract

Alkyltransferase-like proteins (ATLs) share functional motifs with the cancer chemotherapy target O⁶-alkylguanine-DNA alkyltransferase (AGT) and paradoxically protect cells from the biological effects of DNA alkylation damage, despite lacking the reactive cysteine and alkyltransferase activity of AGT. Here we determine Schizosaccharomyces pombe ATL structures without and with damaged DNA containing the endogenous lesion O⁶-methylguanine or cigarette-smoke-derived O⁶-4-(3-pyridyl)-4-oxobutylguanine. These results reveal non-enzymatic DNA nucleotide flipping plus increased DNA distortion and binding pocket size compared to AGT. Our analysis of lesion-binding site conservation identifies new ATLs in sea anemone and ancestral archaea, indicating that ATL interactions are ancestral to present-day repair pathways in all domains of life. Genetic connections to mammalian XPG (also known as ERCC5) and ERCC1 in S. pombe homologues Rad13 and Swi10 and biochemical interactions with Escherichia coli UvrA and UvrC combined with structural results reveal that ATLs sculpt alkylated DNA to create a genetic and structural intersection of base damage processing with nucleotide excision repair.

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