PAHs对抗生素抗性质粒的物理性损伤及机制

Polycyclic aromatic hydrocarbons (PAHs) and antibiotic resistance plasmids (Arps) in environments pose a serious threat to human health and ecological security. PAHs-Arps combined pollution has aroused much public concern. It is necessary to understand the fundamental rules for PAHs-Arps interaction prior to assessing the underlying environmental ecological risks for PAHs-Arps combined pollution. Previous reports have shown that PAHs have an influence on hydrogen bonds, hydrophobic effect between bases and aromatic rings, and Van der Wals forces between bases, physically damaging the DNA structure. But little is known about the physical damage of Arps induced by PAHs in the background of PAHs-Arps combined pollution. In the present study, PAHs and Arps species frequently detected in environments are selected as target pollutants. Research methods including the micro titration, and genetic manipulation etc. and analysis technologies including spectroscopy, computational chemistry, and atomic force microscope are employed in the study to: (1) systematically reveal the fundamental rules of PAHs-induced physical damage to Arps based on the interaction between PAHs and Arps; (2) study the transcription and expression of resistance gene in Arps, identify the relationship between the extent of damage and gene expression level, evaluate the biological effects of damage, and comprehensively expound the mechanisms of PAH-induced physical damages to Arps. It is expected that the information obtained from the study may provide a theoretical foundation for revealing the principle of PAHs-Arps combined pollution, which has great significance for controlling the combined pollution and protecting ecological security in region.

多环芳烃（PAHs）与抗生素抗性质粒（Arps）常共存于环境中，危害人群健康和生态安全，PAHs-Arps复合污染的风险已引起学界关注。弄清PAHs与Arps间的交互作用是阐释其复合污染的前提。已证实，PAHs能影响普通DNA内氢键、碱基对芳香环堆积力、碱基间范德华力，使其发生物理性损伤。然而迄今，关于复合污染背景下PAHs对Arps的物理损伤，相关研究鲜有报道。本项目以常见PAHs和Arps为目标污染物，拟采用微滴定、基因操作等试验方法，利用谱学、计算化学、原子力学等分析技术，在弄清两者互作方式的基础上，系统地揭示PAHs对Arps的物理性损伤规律；研究Arps内抗性基因的转录与表达，找出其损伤程度与基因表达水平间内在联系，评估其损伤的生物学效应，综合性地阐释Arps损伤的机制。成果可为深入揭示PAHs-Arps复合污染原理提供理论基础，对防治其复合污染、保障生态安全具有重要意义。

多环芳烃（PAHs）与抗生素抗性质粒（Arps）常共存于环境中，危害人群健康和生态安全，弄清PAHs与Arps间的相互作用是阐释其复合污染的前提。本项目利用谱学、计算化学、原子力学等分析技术，系统地揭示PAHs对Arps的物理性损伤规律。首先，通过使用固相萃取和计算化学的方法，研究了模型化合物菲与四种基因碱基的相互作用规律，揭示了它们之间的分子驱动机制。在pH7.0条件下，腺嘌呤与菲间的引力较弱，而鸟嘌呤、胞嘧啶和胸腺嘧啶对菲的线性系数约等于1.0，属于分配作用，亲和力较高。酸性条件有利于腺嘌呤对菲的结合（分配，1/n = 1），但不利于胸腺嘧啶与菲的结合。中性或碱性条件下，由于胞嘧啶和胸腺嘧啶存在羰基取代基，作为电子给予体含氧嘧啶脱质子化，表面负电性增强，则整个分子极化减弱，平面л电子更易于与菲弧角位碳原子发生“吸电子-赠电子”作用，亲和作用增强。红外光谱和弱力等值面分析表明，鸟嘌呤和腺嘌呤通过两组CN环（CN六环和五环）同时与菲的两个毗邻苯核发生л-л相互作用；而两类嘧啶仅通过分子平面与菲弧角位置的碳原子发生“赠电子-吸电子”静电引力作用。能量计算的结果表明基因碱基作用位点的预测结果是可信的，属于自发的非共价键成因的物理结合过程。其次，我们进一步探讨了广泛存在于环境中、具有代表性的多环芳烃菲、芘和苯并苝与质粒pUC19的相互作用规律。结果表明，具小分子尺寸的菲更易于与Arps形成松散的团状“Arps-PAHs”复合物，造成Arps内部抗生素抗性基因转换效率的降低。体外转录实验表明，基因转换效率的降低是由于PAHs抑制剂抑制了抗生素抗性基因DNA片段到RNA的转录。荧光微滴定、红外光谱和理论计算模型的综合研究显示，Arps内的腺嘌呤与小分子尺寸的菲和芘之间拥有更强的结合能力；其引力依赖于л-л引力作用。其结合能的变化表明，构建的CT碱基模型和小分子芳香烃之间主要通过非共价的物理吸附相结合。该研究将为深入揭示芳香烃的基因污染提供理论基础，对于防治芳香烃污染、保障生态安全有重要意义。