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 strains. The 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 -diethyl thiophosphate ester (DETP) was used, the reaction rate was determined to be 4.3 ± 0.5 M s in the first-order for both phosphorothioate and peroxynitrite at 35 °C and pH of 8.0. The IC values of phosphorothioate dinucleotides are dramatically increased by 400-700-fold compared to DETP. The SH/OH Yin-Yang mechanism rationalizes the DNA self-defense against PN-mediated oxidative stress at the extra bioenergetic cost of DNA modification.
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