AP站点
化学
DNA
组蛋白
DNA修复
DNA连接酶
组蛋白H4
分子生物学
生物化学
生物
作者
Ying Peng,Xiaoying Wei,Kun Yang
标识
DOI:10.1021/acs.bioconjchem.3c00156
摘要
Histones catalyze the DNA strand incision at apurinic/apyrimidinic (AP) sites accompanied by formation of reversible but long-lived DNA–protein cross-links (DPCs) at 3′-DNA termini within single-strand breaks. These DPCs need to be removed because 3′-hydroxyl is required for gap-filling DNA repair synthesis but are challenging to study because of their reversible nature. Here we report a chemical approach to synthesize stable and site-specific 3′-histone-DPCs and their repair by three nucleases, human AP endonuclease 1, tyrosyl-DNA phosphodiesterase 1, and three-prime repair exonuclease 1. Our method employs oxime ligation to install an alkyne to 3′-DNA terminus, genetic incorporation of an azidohomoalanine to histone H4 at a defined position, and click reaction to conjugate DNA to H4 site-specifically. Using these model DPC substrates, we demonstrated that the DPC repair efficiency is highly affected by the local protein environment, and prior DPC proteolysis facilitates the repair.
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