Visualizing the coordination of apurinic/apyrimidinic endonuclease (APE1) and DNA polymerase β during base excision repair

Base excision repair (BER) is carried out by a series of pro-teins that function in a step-by-step process to identify, remove, and replace DNA damage. During BER, the DNA transitions through various intermediate states as it is processed by each DNA repair enzyme. Left unrepaired, these BER intermediates can transition into double-stranded DNA breaks and promote genome instability. Previous studies have proposed a short-lived complex consisting of the BER intermediate, the incoming enzyme, and the outgoing enzyme at each step of the BER pathway to protect the BER intermediate. The transfer of BER intermediates between enzymes, known as BER coordi-nation or substrate channeling, remains poorly understood. Here, we utilize single-molecule total internal reflection fluo-rescence microscopy to investigate the mechanism of BER co-ordination between apurinic/apyrimidinic endonuclease 1 (APE1) and DNA polymerase beta (Pol beta). When preformed complexes of APE1 and the incised abasic site product (APE1 product and Pol beta substrate) were subsequently bound by Pol beta, the Pol beta enzyme dissociated shortly after binding in most of the observations. In the events where Pol beta binding was fol-lowed by APE1 dissociation during substrate channeling, Pol beta remained bound for a longer period of time to allow disasso-ciation of APE1. Our results indicate that transfer of the BER intermediate from APE1 to Pol beta during BER is dependent on the dissociation kinetics of APE1 and the duration of the ternary complex on the incised abasic site.

Automated summary

Base excision repair (BER) is a multi-step process that involves the coordinated action of multiple proteins to identify, remove, and replace DNA damage. Understanding the transfer of BER intermediates between enzymes, a process known as BER coordination or substrate channeling, is crucial in maintaining genome stability. In this study, single-molecule microscopy was used to investigate the mechanism of BER coordination between APE1 and Pol β. The results show that the transfer of BER intermediate from APE1 to Pol β is dependent on the dissociation kinetics of APE1 and the duration of the ternary complex on the incised abasic site.