9DP3

APE1 N174D Product Complex with Abasic DNA

9DP3 の概要

• エントリーDOI: 10.2210/pdb9dp3/pdb
• 分子名称: DNA repair nuclease/redox regulator APEX1, mitochondrial, DNA (5'-D(P*(3DR)P*CP*GP*AP*CP*GP*GP*AP*TP*CP*C)-3'), DNA (5'-D(*GP*CP*TP*GP*AP*TP*GP*CP*GP*C)-3'), ... (6 entities in total)
• 機能のキーワード: ape1, apex1, endonuclease, dna repair, hydrolase-dna complex, hydrolase/dna
• 由来する生物種: Homo sapiens (human); 詳細
• タンパク質・核酸の鎖数: 5
• 化学式量合計: 75147.24

構造登録者

Hoitsma, N.M.,DeHart, K.D.,Freudenthal, B.D. (登録日: 2024-09-20, 公開日: 2024-10-09, 最終更新日: 2025-10-29)

主引用文献

DeHart, K.M.,Hoitsma, N.M.,Thompson, S.H.,Borin, V.A.,Agarwal, P.K.,Freudenthal, B.D.
APE1 active site residue Asn174 stabilizes the AP site and is essential for catalysis.
J.Biol.Chem., 301:110655-110655, 2025

PubMed Abstract: Apurinic/apyrimidinic (AP) sites are common and highly mutagenic DNA lesions that can arise spontaneously or as intermediates during base excision repair. The enzyme apurinic/apyrimidinic endonuclease 1 (APE1) initiates repair of AP sites by cleaving the DNA backbone at the AP site via its endonuclease activity. Here, we investigated the functional role of the APE1 active site residue N174 that contacts the AP site during catalysis. To do so, we rationally designed four APE1 mutants (N174A, N174D, N174Q, and N174K) that altered the hydrogen bonding potential, electronegativity, and side chain length of residue 174. Kinetic analysis determined that AP site cleavage was impaired in our APE1, APE1, and APE1 mutants, but not in our APE1 mutant. Using X-ray crystallography and molecular dynamic simulations of our APE1 mutants, we attributed the poor cleavage of our APE1 and APE1 mutants to lack of hydrogen bonding between residue 174 and the AP site. From these experiments, we also discovered that the altered charge of residue 174 in APE1 and APE1 mutants contributed to impaired cleavage. From analysis of our mutants, we determined that in APE1, N174 stabilizes AP DNA within the APE1 active site via hydrogen bonding promoting cleavage of AP sites. We also found that the neutral charge of N174 in APE1 contributes to an optimal electrostatic environment during the APE1 endonuclease reaction resulting in efficient AP site cleavage. Cumulatively, we demonstrate the importance of N174 in APE1's function and provide new insights into the molecular mechanism by which APE1 processes AP sites during DNA repair.

PubMed: 40902980
DOI: 10.1016/j.jbc.2025.110655

実験手法

X-RAY DIFFRACTION (2.1 Å)