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In response to oxidative damage, base excision repair (BER) enzymes perturb the structural equilibrium of the VEGF promoter between B‐form and G4 DNA conformations, resulting in epigenetic‐like modifications of gene expression. However, the mechanistic details remain enigmatic, including the activity and coordination of BER enzymes on the damaged G4 promoter. To address this, we investigated the ability of each BER factor to conduct its repair activity on VEGF promoter G4 DNA substrates by employing pre‐steady‐state kinetics assays and in vitro coupled BER assays. OGG1 was able to initiate BER on double‐stranded VEGF promoter G4 DNA substrates. Moreover, pre‐steady‐state kinetics revealed that compared to B‐form DNA, APE1 repair activity on the G4 was decreased ~two‐fold and is the result of slower product release as opposed to inefficient strand cleavage. Interestingly, Pol β performs multiple insertions on G4 substrates via strand displacement DNA synthesis in contrast to a single insertion on B‐form DNA. The multiple insertions inhibit ligation of the Pol β products, and hence BER is not completed on the VEGF G4 promoter substrates through canonical short‐patch BER. Instead, repair requires the long‐patch BER flap‐endonuclease activity of FEN1 in response to the multiple insertions by Pol β prior to ligation. Because the BER proteins and their repair activities are a key part of the VEGF transcriptional enhancement in response to oxidative DNA damage of the G4 VEGF promoter, the new insights reported here on BER activity in the context of this promoter are relevant toward understanding the mechanism of transcriptional regulation. [ABSTRACT FROM AUTHOR]

DNA is susceptible to a range of chemical modifications, with one of the most frequent lesions being apurinic/apyrimidinic (AP) sites. AP sites arise due to damage-induced (e.g. alkylation) or spontaneous hydrolysis of the N-glycosidic bond that links the base to the sugar moiety of the phosphodiester backbone, or through the enzymatic activity of DNA glycosylases, which release inappropriate bases as part of the base excision repair (BER) response. Unrepaired AP sites, which lack instructional information, have the potential to cause mutagenesis or to arrest progressing DNA or RNA polymerases, potentially causing outcomes such as cellular transformation, senescence or death. The predominant enzyme in humans responsible for repairing AP lesions is AP endonuclease 1 (APE1). Besides being a powerful AP endonuclease, APE1 possesses additional DNA repair activities, such as 3′–5′ exonuclease, 3′-phophodiesterase and nucleotide incision repair. In addition, APE1 has been shown to stimulate the DNA-binding activity of a number of transcription factors through its 'REF1' function, thereby regulating gene expression. In this article, we review the structural and biochemical features of this multifunctional protein, while reporting on new structures of the APE1 variants Cys65Ala and Lys98Ala. Using a functional complementation approach, we also describe the importance of the repair and REF1 activities in promoting cell survival, including the proposed passing-the-baton coordination in BER. Finally, results are presented indicating a critical role for APE1 nuclease activities in resistance to the genotoxins methyl methanesulphonate and bleomycin, supporting biologically important functions as an AP endonuclease and 3′-phosphodiesterase, respectively. [ABSTRACT FROM AUTHOR]

Reactive oxygen species continuously assault the structure of DNA resulting in oxidation and fragmentation of the nucleobases. Both oxidative DNA damage itself and its repair mediate the progression of many prevalent human maladies. The major pathway tasked with removal of oxidative DNA damage, and hence maintaining genomic integrity, is base excision repair (BER). The aphorism that structure often dictates function has proven true, as numerous recent structural biology studies have aided in clarifying the molecular mechanisms used by key BER enzymes during the repair of damaged DNA. This review focuses on the mechanistic details of the individual BER enzymes and the association of these enzymes during the development and progression of human diseases, including cancer and neurological diseases. Expanding on these structural and biochemical studies to further clarify still elusive BER mechanisms, and focusing our efforts toward gaining an improved appreciation of how these enzymes form co-complexes to facilitate DNA repair is a crucial next step toward understanding how BER contributes to human maladies and how it can be manipulated to alter patient outcomes.