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African swine fever virus (ASFV) is highly contagious and can cause lethal disease in pigs. ASFV is primarily ****ted in the cytoplasm of pig macrophages, which is oxidative and caused constant damage to ASFV genome. ASFV AP endonuclease (AsfvAP) catalyzes DNA cleavage reaction at the abasic site and is a key enzyme of ASFV base excision repair (BER) system. Although it plays an essential role in ASFV survival in host cells, the basis underlying substrate binding and cleavage by AsfvAP remains unclear. Here, we reported the structura