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Title: Transient kinetic analysis of oxidative dealkylation by the direct reversal DNA repair enzyme AlkB
AlkB is a bacterial Fe(II)– and 2-oxoglutarate–dependent dioxygenase that repairs a wide range of alkylated nucleobases in DNA and RNA as part of the adaptive response to exogenous nucleic acid–alkylating agents. Although there has been longstanding interest in the structure and specificity of Escherichia coli AlkB and its homologs, difficulties in assaying their repair activities have limited our understanding of their substrate specificities and kinetic mechanisms. Here, we used quantitative kinetic approaches to determine the transient kinetics of recognition and repair of alkylated DNA by AlkB. These experiments revealed that AlkB is a much faster alkylation repair enzyme than previously reported and that it is significantly faster than DNA repair glycosylases that recognize and excise some of the same base lesions. We observed that whereas 1, N 6 -ethenoadenine can be repaired by AlkB with similar efficiencies in both single- and double-stranded DNA, 1-methyladenine is preferentially repaired in single-stranded DNA. Our results lay the groundwork for future studies of AlkB and its human homologs ALKBH2 and ALKBH3.  more » « less
Award ID(s):
1615586
PAR ID:
10156610
Author(s) / Creator(s):
; ;
Date Published:
Journal Name:
Journal of Biological Chemistry
Volume:
295
Issue:
21
ISSN:
0021-9258
Page Range / eLocation ID:
7317 to 7326
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
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