The influence of XRCC1 in the early stage of BER initiated by MPG.

Base excision repair (BER) is one of the dominant pathways to repair altered DNA bases, and is well conserved from bacteria to mammals. Single nucleotide BER (SN-BER) is a simple pathway to remove a single nucleotide. We have investigated several protein–protein interactions and their coordination in SN-BER initiated by methylpurine-DNA glycosylase (MPG). MPG recognizes and excises damaged purines, such as hypoxantin or methylated bases, from DNA. With activity assays using radio-labeled duplex oligonucleotide substrates, we concluded that the downstream enzymes increase the upstream enzyme activity in SN-BER. For example, AP endonuclease 1 (APE1) increased MPG activity, and DNA polymerase β increased MPG and APE1 activity. Since the stimulations resulted from increases of kcat values, a downstream enzyme would accelerate the disassociation of the upstream enzyme from the product. Intracellular X-ray repair cross complementing protein 1 (XRCC1) is thought to make complexes with 80 percent of DNA Ligase 3 (LIG3) molecules under physiological conditions, and interacts with almost all of BER enzymes to stabilize the configuration. In the presence of recombinant XRCC1, kcat/Km value of MPG and APE1 activity increased 1.8-fold and 7.1-fold, respectively, suggesting the involvement in a repair complex during the early step of SN-BER pathway. The studies on the effects of XRCC1-LIG3 complex to MPG and APE1 activities are in progress.