Abstract

Efficient recognition of DNA lesions such as apurinic/apyrimidinic (AP) sites is essential for maintaining genome stability. Apurinic/apyrimidinic endonuclease 1 (APE1) is the primary eukaryotic AP endonuclease, yet how it identifies rare lesions among vast stretches of undamaged DNA remains incompletely understood. Using single-molecule imaging combined with molecular dynamics simulations, we reveal that APE1 employs a distinctive dual mechanism to search DNA damage. First, Mg2+ coordination at the active site neutralizes clustered negative charges, stabilizing electrostatic contacts during scanning. Second, its N-terminal intrinsically disordered region (IDR)—a feature conserved only in eukaryotic homologs but absent in prokaryotic ExoIII—not only interacts with DNA through transient IDR contacts but also engages continuous interactions via the unprecedented Arg177 residue within the structured nuclease domain, thereby prolonging residence time and enabling long-range diffusion. Together, these two modules synergize to promote a sliding-based search strategy tailored to the complexity of eukaryotic genomes. Consistent with this model, IDR deletion restricts APE1 to 3D collisions, whereas IDR duplication enhances 1D scanning. Thus, APE1 exemplifies how structural disorder and metal-ion coordination integrates to enable long-range lesion recognition, highlighting an evolutionary innovation in eukaryotic DNA repair.

A research team, affiliated with UNIST, has uncovered the molecular details behind how the DNA repair enzyme APE1 locates damaged sites within the genome with remarkable speed. Their findingsshed light on a sophisticated search mechanism that enables cells to maintain genetic stability.

DNA is continually damaged by environmental factors and cellular processes. Among these, apurinic/apyrimidinic (AP) sites—where a base is missing—pose a serious threat if left unrepaired. Detecting these rare lesions within billions of undamaged bases is like searching for a needle in a haystack. 

To understand how the repair enzyme APE1 efficiently locataes these sites, Professor Ja Yil Lee from the Department of Biological Sciences, in collaboration with Professor Gwangrog Lee at KAIST and Professor Jejoong Yoo at Sungkyunkwan University (SKKU), conducted detailed research. They employed advanced single-molecule techniques—such as FRET assays and DNA curtain methods—along with molecular dynamics simulations to observe APE1's behavior in real time.

Their findings show that APE1 does not scan DNA randomly. Instead, it slides along the strand in a one-dimensional manner, rapidly searching for damage. This sliding mechanism resembles a smart robot navigating a complex underground pipeline network, efficiently pinpointing leaks rather than wandering aimlessly.

A key discovery involves APE1's flexible, disordered region—an intrinsically disordered segment that functions as a molecular hook. This region facilitates the enzyme's attachment to DNA during its search. Removal of this segment reduced APE1's damage-finding ability by more than fivefold.

The study also highlights the role of magnesium ions (Mg²⁺). Beyond their known function as cofactors, magnesium ions stabilize APE1's interaction with DNA, enhancing its mobility and search efficiency.

Professor Gwangrok Lee of KAIST remarked that these findings shed light on how flexible protein regions and metal ions work together to enable rapid damage detection, which could inform the development of new cancer therapies.

Professor Ja Yil Lee added, “Understanding this mechanism opens new avenues for designing treatments that modulate DNA repair, with potential applications in cancer and aging.”

Their findings have been published in Nucleic Acids Research on May 14, 2026. The study was supported by KAIST Grand Challenge 30 Project (KC30), the National Research Foundation of the Korea (NRF), the Institute for Basic Science (IBS), and the Institute of Information & communications Technology Planning & Evaluation (IITP).

Journal Reference

Donghun Lee, Subin Kim, Gyeongpil Jo, et al ., " APE1 coordinates its disordered region and metal cofactors to drive genome surveillance,"   NAR , (2026).