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Issue 39, 2017
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Structural basis of interstrand cross-link repair by O6-alkylguanine DNA alkyltransferase

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Abstract

DNA interstrand cross-links (ICL) are among the most cytotoxic lesions found in biological systems. O6-Alkylguanine DNA alkyltransferases (AGTs) are capable of removing alkylation damage from the O6-atom of 2′-deoxyguanosine and the O4-atom of thymidine. Human AGT (hAGT) has demonstrated the ability to repair an interstrand cross-linked duplex where two O6-atoms of 2′-deoxyguanosine were tethered by a butylene (XLGG4) or heptylene (XLGG7) linkage. However, the analogous ICL between the O4-atoms of thymidine was found to evade repair. ICL duplexes connecting the O4-atoms of 2′-deoxyuridine by a butylene (XLUU4) or heptylene (XLUU7) linkage have been prepared to examine the influence of the C5-methyl group on AGT-mediated repair. Both XLUU4 and XLUU7 were refractory to repair by human and E. coli (OGT and Ada-C) AGTs with comparably low μM dissociation constants for 2 : 1 or 4 : 1 AGT/DNA stoichiometries. The solution structures of two heptylene linked DNA duplexes (CGAAAYTTTCG)2, XLUU7 (Y = dU) and XLGG7 (Y = dG), were solved and the global structures were virtually identical with a RMSD of 1.22 Å. The ICL was found to reside in the major groove for both duplexes. The linkage adopts an E conformation about the C4–O4 bond for XLUU7 whereas a Z conformation about the C6–O6 bond was observed for XLGG7. This E versus Z conformation may partially account for hAGTs discrimination towards the repair of these ICL, supported by the crystal structures of hAGT with various substrates which have been observed to adopt a Z conformation. In addition, a higher mobility at the ICL site for XLUU7 is observed relative to XLGG7 that may play a role in repair by hAGT. Taken together, these findings provide insights on the AGT-mediated repair of cytotoxic ICL in terms of its processing capability and substrate specificity.

Graphical abstract: Structural basis of interstrand cross-link repair by O6-alkylguanine DNA alkyltransferase

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Supplementary files

Article information


Submitted
22 Aug 2017
Accepted
14 Sep 2017
First published
15 Sep 2017

Org. Biomol. Chem., 2017,15, 8361-8370
Article type
Paper

Structural basis of interstrand cross-link repair by O6-alkylguanine DNA alkyltransferase

A. Y. Denisov, F. P. McManus, D. K. O'Flaherty, A. M. Noronha and C. J. Wilds, Org. Biomol. Chem., 2017, 15, 8361
DOI: 10.1039/C7OB02093G

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