DNA phosphorothioation (PT) exists in many pathogenic bacteria; however, the mechanism of PT-DNA resistance to the immune response is unclear. In this work, we meticulously investigated the peroxynitrite (PN) tolerance using PT-bioengineered E. coli strains. The in vivo experiment confirms that the S⁺ strain survives better than the S^– strain under moderately oxidative stress. The LCMS, IC, and GCMS experiments demonstrated that phosphorothioate partially converted to phosphate, and the byproduct included sulfate and elemental sulfur. When O,O-diethyl thiophosphate ester (DETP) was used, the reaction rate k₁ was determined to be 4.3 ± 0.5 M^–1 s^–1 in the first-order for both phosphorothioate and peroxynitrite at 35 °C and pH of 8.0. The IC₅₀ values of phosphorothioate dinucleotides are dramatically increased by 400–700-fold compared to DETP. The SH/OH Yin-Yang mechanism rationalizes the in situ DNA self-defense against PN-mediated oxidative stress at the extra bioenergetic cost of DNA modification.

中文翻译：

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过亚硝酸盐介导的氧化应激下硫代磷酸化DNA的防御机制。

DNA硫代磷酸化（PT）存在于许多致病细菌中。然而，PT-DNA抵抗免疫反应的机制尚不清楚。在这项工作中，我们使用PT生物工程化的大肠杆菌菌株精心研究过亚硝酸盐（PN）的耐受性。在体内实验确认了在S ⁺应变幸存比S更好^-适度氧化应激下的应变。LCMS，IC和GCMS实验表明，硫代磷酸酯部分转化为磷酸盐，副产物包括硫酸盐和元素硫。当使用O，O-二乙基硫代磷酸酯（DETP）时，反应速率k ₁被确定为4.3±0.5 M ^–1 s在35°C和pH值为8.0时，硫代磷酸酯和过氧亚硝酸盐的一级为^–1。与DETP相比，硫代磷酸二核苷酸的IC ₅₀值显着提高了400-700倍。SH / OH阴阳机制以DNA修饰所需的额外生物能成本合理化了针对PN介导的氧化应激的原位DNA自防御。