African swine fever virus (ASFV) is highly contagious and can cause lethal disease in pigs. ASFV is primarily replicated 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 structural and functional studies of AsfvAP, showing that AsfvAP adopts a novel DNA-binding mode distinct from other APs. AsfvAP possesses many unique structural features, including one narrower nucleotide-binding pocket at the active site, the C16–C20 disulfide bond-containing region, and histidine-rich loop. As indicated by our mutagenesis, in vitro binding and cleavage assays, these features are important for AsfvAP to suit the acidic and oxidative environment. Owing to their functional importance, these unique features could serve as targets for designing small molecule inhibitors that could disrupt the repair process of ASFV genome and help fight against this deadly virus in the future.

非洲猪瘟病毒（ASFV）具有高度传染性，可导致猪致命。ASFV主要在猪巨噬细胞的细胞质中复制，巨噬细胞具有氧化性，对ASFV基因组造成持续损伤。ASFV AP 核酸内切酶（Asfv AP）催化脱碱基位点的 DNA 裂解反应，是 ASFV 碱基切除修复（BER）系统的关键酶。尽管它在 ASFV 在宿主细胞中的存活中发挥着重要作用，但Asfv AP与底物结合和裂解的基础仍不清楚。在这里，我们报道了Asfv AP的结构和功能研究，表明Asfv AP采用了不同于其他AP的新型DNA结合模式。Asfv AP 具有许多独特的结构特征，包括活性位点上较窄的核苷酸结合袋、含有 C16-C20 二硫键的区域以及富含组氨酸的环。正如我们的诱变、体外结合和裂解测定所示，这些特征对于Asfv AP 适应酸性和氧化环境非常重要。由于其功能重要性，这些独特的特征可以作为设计小分子抑制剂的目标，这些抑制剂可以破坏 ASFV 基因组的修复过程，并有助于在未来对抗这种致命病毒。