Synthesis and Excision Repair of Site-Specific 3′-End DNA-Histone Cross-Links Derived from Abasic Sites

Histonescatalyze the DNA strand incision at apurinic/apyrimidinic(AP) sites accompanied by formation of reversible but long-lived DNA-proteincross-links (DPCs) at 3 & PRIME;-DNA termini within single-strand breaks.These DPCs need to be removed because 3 & PRIME;-hydroxyl is requiredfor gap-filling DNA repair synthesis but are challenging to studybecause of their reversible nature. Here we report a chemical approachto synthesize stable and site-specific 3 & PRIME;-histone-DPCs andtheir repair by three nucleases, human AP endonuclease 1, tyrosyl-DNAphosphodiesterase 1, and three-prime repair exonuclease 1. Our methodemploys oxime ligation to install an alkyne to 3 & PRIME;-DNA terminus,genetic incorporation of an azidohomoalanine to histone H4 at a definedposition, and click reaction to conjugate DNA to H4 site-specifically.Using these model DPC substrates, we demonstrated that the DPC repairefficiency is highly affected by the local protein environment, andprior DPC proteolysis facilitates the repair.

Automated summary

Histones catalyze DNA strand incision at AP sites and form reversible but long-lived DPCs at 3'-DNA termini within single-strand breaks. This study developed a chemical approach to synthesize stable and site-specific 3'-histone-DPCs and their repair by nucleases. The efficiency of DPC repair is influenced by the local protein environment, and previous DPC proteolysis facilitates the repair.