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Title: The DNA glycosylase AlkD uses a non-base-flipping mechanism to excise bulky lesions

Journal Article · · Nature (London)
DOI:https://doi.org/10.1038/nature15728· OSTI ID:1229895
 [1];  [1];  [1];  [2];  [2];  [3];  [1]
  1. Vanderbilt Univ., Nashville, TN (United States). Dept. of Biological Sciences. Center for Structural Biology
  2. Univ. of California, Davis, CA (United States). Dept. of Chemistry
  3. Toyama Prefectural Univ., Imizu (Japan). Biotechnology Research Center
+ Show Author Affiliations

Threats to genomic integrity arising from DNA damage are mitigated by DNA glycosylases, which initiate the base excision repair pathway by locating and excising aberrant nucleobases. How these enzymes find small modifications within the genome is a current area of intensive research. A hallmark of these and other DNA repair enzymes is their use of base flipping to sequester modified nucleotides from the DNA helix and into an active site pocket. Consequently, base flipping is generally regarded as an essential aspect of lesion recognition and a necessary precursor to base excision. In this paper, we present the first, to our knowledge, DNA glycosylase mechanism that does not require base flipping for either binding or catalysis. Using the DNA glycosylase AlkD from Bacillus cereus, we crystallographically monitored excision of an alkylpurine substrate as a function of time, and reconstructed the steps along the reaction coordinate through structures representing substrate, intermediate and product complexes. Instead of directly interacting with the damaged nucleobase, AlkD recognizes aberrant base pairs through interactions with the phosphoribose backbone, while the lesion remains stacked in the DNA duplex. Quantum mechanical calculations revealed that these contacts include catalytic charge–dipole and CH–π interactions that preferentially stabilize the transition state. We show in vitro and in vivo how this unique means of recognition and catalysis enables AlkD to repair large adducts formed by yatakemycin, a member of the duocarmycin family of antimicrobial natural products exploited in bacterial warfare and chemotherapeutic trials. Bulky adducts of this or any type are not excised by DNA glycosylases that use a traditional base-flipping mechanism. Finally and hence, these findings represent a new model for DNA repair and provide insights into catalysis of base excision.

Research Organization:
Vanderbilt Univ., Nashville, TN (United States)
Sponsoring Organization:
USDOE Office of Science (SC); National Science Foundation (NSF); National Inst. of Health (NIH) (United States); Michigan Economic Development Corporation (United States); Michigan Technology Tri-Corridor (United States); Vanderbilt Univ. (United States)
Contributing Organization:
Univ. of California, Davis, CA (United States); Toyama Prefectural Univ., Imizu (Japan)
Grant/Contract Number:
AC02-06CH11357; MCB-1122098; MCB-1517695; R01ES019625; R01CA067985; S10RR026915; 085P1000817; T32ES07028
OSTI ID:
1229895
Journal Information:
Nature (London), Vol. 527, Issue 7577; ISSN 0028-0836
Publisher:
Nature Publishing GroupCopyright Statement
Country of Publication:
United States
Language:
ENGLISH
Citation Metrics:
Cited by: 45 works
Citation information provided by
Web of Science

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Cited By (5)

Toxicity and repair of DNA adducts produced by the natural product yatakemycin journal July 2017
Evolutionary diversity and novelty of DNA repair genes in asexual Bdelloid rotifers journal November 2018
A vitamin-C-derived DNA modification catalysed by an algal TET homologue journal May 2019
Insights into conformational changes in AlkD bound to DNA with a yatakemycin adduct from computational simulations journal May 2018
Combining crystallographic and quantum chemical data to understand DNA-protein π-interactions in nature journal April 2017

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