GO system is part of base excision DNA repair and is required for the correct repair of 8-oxoguanine (8-oxoG), one of the most abundant oxidative lesions. Due to the ability of 8-oxoG to mispair with A, this base is highly mutagenic, and its repair requires two enzymes: Fpg that removes 8-oxoG from 8-oxoG:C pairs, and MutY that excises the normal A from 8-oxoG:A mispairs. Here we characterize the properties of putative GO system DNA glycosylases from Staphylococcus aureus, an important human opportunistic pathogen that causes hospital infections and presents a serious health concern due to quick spread of antibiotic-resistant strains. In addition to Fpg and MutY from the reference NCTC 8325 strain (SauFpg1 and SauMutY), we have also studied an Fpg homolog from a multidrug-resistant C0673 isolate (SauFpg2), which is different from SauFpg1 in its sequence. Both SauFpg enzymes showed the highest activity at pH 7.0–9.0 and NaCl concentrations 25–75 mM (SauFpg1) or 50–100 mM (SauFpg2), whereas SauMutY was active at a broad pH range and had a salt optimum at ∼75 mM NaCl. Both SauFpg1 and SauFpg2 bound and cleaved duplexes containing 8-oxoG, 5-hydroxyuracil, 5,6-dihydrouracil or apurinic/apyrimidinic site paired with C, T, or G, but not with A. For SauFpg1 and SauFpg2, 8-oxoG was the best substrate tested, and 5,6-dihydrouracil was the worst one. SauMutY efficiently excised adenine from duplex substrates containing A:8-oxoG or A:G pairs. SauFpg enzymes were readily trapped on DNA by NaBH₄ treatment, indicating formation of a Schiff base reaction intermediate. Surprisingly, SauMutY was also trapped significantly better than its E. coli homolog. All three S. aureus GO glycosylases drastically reduced spontaneous mutagenesis when expressed in an fpg mutY E. coli double mutant. Overall, we conclude that S. aureus possesses an active GO system, which could possibly be targeted for sensitization of this pathogen to oxidative stress.

GO 系统是碱基切除 DNA 修复的一部分，是正确修复 8-氧鸟嘌呤 (8-oxoG)（最丰富的氧化损伤之一）所必需的。由于 8-oxoG 与 A 错配的能力，该碱基具有高度致突变性，其修复需要两种酶：从 8-oxoG:C 对中去除 8-oxoG 的 Fpg，以及从 8-oxoG:C 对中去除正常 A 的 MutY oxoG：A错配。在这里，我们表征了来自金黄色葡萄球菌的推定 GO 系统 DNA 糖基化酶的特性，一种重要的人类机会性病原体，会导致医院感染并由于抗生素耐药菌株的快速传播而引起严重的健康问题。除了来自参考 NCTC 8325 菌株（SauFpg1 和 SauMutY）的 Fpg 和 MutY，我们还研究了来自多药耐药 C0673 分离株（SauFpg2）的 Fpg 同源物，其序列与 SauFpg1 不同。两种 SauFpg 酶在 pH 7.0–9.0 和 NaCl 浓度 25–75 mM (SauFpg1) 或 50–100 mM (SauFpg2) 下均表现出最高活性，而 SauMutY 在较宽的 pH 范围内具有活性，并且在约 75 mM NaCl 下具有最佳盐分. SauFpg1 和 SauFpg2 都结合和切割含有 8-oxoG、5-羟基尿嘧啶、5,6-二氢尿嘧啶或无嘌呤/无嘧啶位点的双链体，与 C、T 或 G 配对，但不与 A 配对。对于 SauFpg1 和 SauFpg2，8-oxoG 是测试的最佳基材，和 5, 6-二氢尿嘧啶是最差的一种。SauMutY 从含有 A:8-oxoG 或 A:G 对的双链底物中有效切除腺嘌呤。SauFpg 酶很容易被 NaBH 捕获在 DNA 上₄处理，表明形成席夫碱反应中间体。令人惊讶的是，SauMutY 的捕获效果也明显优于其大肠杆菌同系物。当在fpg mutY 大肠杆菌双突变体中表达时，所有三种金黄色葡萄球菌GO 糖基化酶都显着降低了自发诱变。总的来说，我们得出结论，金黄色葡萄球菌拥有一个活跃的 GO 系统，它可能是该病原体对氧化应激敏感的目标。