Defining the functional footprint for recognition and repair of deaminated DNA

被引:11
作者
Baldwin, Michael R. [1 ]
O'Brien, Patrick J. [1 ]
机构
[1] Univ Michigan, Dept Biol Chem, Ann Arbor, MI 48109 USA
基金
美国国家卫生研究院;
关键词
HUMAN APURINIC/APYRIMIDINIC ENDONUCLEASE; BASE EXCISION; AP-ENDONUCLEASE; SUBSTRATE-SPECIFICITY; INITIAL STEPS; GLYCOSYLASE; HYPOXANTHINE; BINDING; PROTEIN; OLIGODEOXYRIBONUCLEOTIDES;
D O I
10.1093/nar/gks952
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Spontaneous deamination of DNA is mutagenic, if it is not repaired by the base excision repair (BER) pathway. Crystallographic data suggest that each BER enzyme has a compact DNA binding site. However, these structures lack information about poorly ordered termini, and the energetic contributions of specific protein-DNA contacts cannot be inferred. Furthermore, these structures do not reveal how DNA repair intermediates are passed between enzyme active sites. We used a functional footprinting approach to define the binding sites of the first two enzymes of the human BER pathway for the repair of deaminated purines, alkyladenine DNA glycosylase (AAG) and AP endonuclease (APE1). Although the functional footprint for full-length AAG is explained by crystal structures of truncated AAG, the footprint for full-length APE1 indicates a much larger binding site than is observed in crystal structures. AAG turnover is stimulated in the presence of APE1, indicating rapid exchange of AAG and APE1 at the abasic site produced by the AAG reaction. The coordinated reaction does not require an extended footprint, suggesting that each enzyme engages the site independently. Functional footprinting provides unique information relative to traditional footprinting approaches and is generally applicable to any DNA modifying enzyme or system of enzymes.
引用
收藏
页码:11638 / 11647
页数:10
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