Saccharomyces cerevisiae apurinic/apyrimidinic endonuclease 1 repairs abasic site-mediated DNA-peptide/protein cross-links.

ABSTRACT

Saccharomyces cerevisiae apurinic/apyrimidinic (AP) endonuclease 1 (yApn1) is a key player of the base excision repair pathway. This multifunctional enzyme is an AP endonuclease, 3'-5' exonuclease, 3'-phosphodiesterase, and participates in nucleotide incision repair. To the best of our knowledge, the known substrates of yApn1 are small DNA lesions such as AP sites and 3'-phospho-α,ß-unsaturated aldehyde (3'-PUA). Here, we wish to report in vitro findings that yApn1 repairs bulky DNA-peptide cross-links (DpCs) and DNA-protein cross-links (DPCs) arising from AP sites and 3'-PUA. We chemically synthesized stable and linkage-defined DpCs and DPCs by oxime ligation and reductive amination, respectively. Our steady-state kinetic data showed that yApn1 repairs a 10-mer peptide-conjugated AP site and 3'-PUA with comparable efficiencies to that of processing the unconjugated lesions. We demonstrated that yApn1 is the predominant enzyme that incises AP-DpC in yeast cell extracts. We also demonstrated that yApn1 incises AP-DPCs in a DPC size-dependent manner, and prior DPC proteolysis by trypsin facilitates the repair. We further found that yApn1 removes 3'-PUA-histone DPCs with moderate efficiencies. Together, our results uncovered a novel role of yApn1 in DPC repair, and support the emerging model that proteolysis is required for efficient DPC repair.