PBDEs微生物酶促降解的分子作用机制及其构效关系

Due to the great risks to ecological system, polybrominated diphenyl ethers (PBDEs) have been typically categorized into the lists of POPs. The aerobic biodegradation process of PBDEs was recently proved to be controlled and influenced by the important bph path and further modulated by bphA-encoded enzyme, namely biphenyl dioxygenase (BPDO). However, the intrinsic molecular mechanism of microbial enzymatic degradation has net yet been clearly understood. Based on the biodegradation kinetics of PBDEs with Burkholderia xenovorans LB400, the present project was designed to get insight into the molecular mechanism. With the combination of analysis of hydroxylated intermediate product and computation of electron distribution in frontier molecular orbital by density functional theory, the regioselectivity of hydroxylation sites in biodegradation of PBDEs will be described. The binding interaction between PBDEs and the important subunit of BPDO, viz. BphA1 subunit, can be simulated with the molecular docking technique, molecular dynamics and quantumatic ONIOM method. The distribution of key amino acid residues in active pocket of BPDO and their relative contribution to total energy is thus reflected. Simultaneously, the effect imposed by the binding interaction would also be addressed with the relationship between binding affinity and biodegradation rate constants. The quantitative structure-activity relationships (2D/3D-QSARs) for biodegradation rate constants of PBDEs would be developed with various modelling techniques. With the full consideration of regioselectivity of hydroxylation sites in biodegradation process and the binding interaction between PBDEs and active regions of BPDO, the model analyses would help illuminate the molecular mechanism of enzymatic degradation of PBDEs from the perspective of microcosmic level, as well as the pivotal influential factors upon it. With the help of study mentioned above, it is expected to provide the scientific guides for the screening of PBDEs-degrading bacteria with high efficiency and the practical application in pollution remediation.

多溴联苯醚（PBDEs）属典型持久性有机污染物，已证实其好氧微生物降解过程受bphA基因编码酶-联苯双加氧酶（BPDO）催化调控，然而，有关酶促降解的分子作用机制尚未明确。本项目拟以伯克氏降解菌LB400对PBDEs降解动力学实验为基础，结合羟基化中间产物分析与密度泛函方法对前线分子轨道电子排布计算，揭示PBDEs微生物降解的羟基化区域选择性；采用分子对接、分子动力学和量子化学分层优化方法，模拟研究PBDEs与BPDO关键亚基BphA1的亲合作用，反映活性空穴氨基酸残基分布特征和相对能量贡献，以亲合能与降解速率常数的相关性阐释其对微生物降解性能影响；以多种模型构建技术，发展PBDEs微生物降解速率的定量构效关系（2D/3D-QSARs），基于模型分析，综合微生物降解区域选择性与亲合作用，从微观角度阐明PBDEs酶促降解的分子作用机制及关键影响因子，为高效降解菌选育与污染修复提供科学依据。

多溴联苯醚（PBDEs）是环境中广泛分布且极具生态风险的一类持久性有机污染物，威胁生态系统安全。尽管好氧微生物降解是污染消减的重要途径之一且已证实PBDEs好氧微生物降解受bph基因编码酶催化调控，但有关酶促降解的分子作用机制尚不清晰。本项目拟以密度泛函理论（DFT）计算、Surflex-Dock分子对接和模型发展等技术方法，基于好氧降解菌LB400对不同溴代模式PBDEs的降解动力学过程及共代谢底物、表面活性剂和生物吸附对降解性能的影响研究，揭示好氧微生物降解的热力学机制和羟基化区域选择性，明确降解菌活性酶BphA1活性空穴关键氨基酸残基分布特征及其与PBDEs的亲合作用，阐释PBDEs亲合作用对降解性能的影响，建立与PBDEs结构特征和亲合作用相关联的二维/三维定量构效关系（2D/3D-QSARs）。研究结果表明，1）共代谢底物和表面活性剂对好氧微生物降解具有显著影响。120h内PBDEs去除主要来自好氧降解作用，降解效率为38.4%~100%，降解过程符合一级反应动力学（R2在0.814~0.991）。随着PBDEs溴代数目增加，降解平衡时间延长且降解速率下降。2）短期内LB400降解菌对BDE-47的去除主要归因于生物吸附作用，随培养时间延长，降解作用所占比例增加并占据主导。3）羟基化反应是PBDEs微生物降解反应的速控步骤，活性氧自由基如•OH更倾向于攻击苯环的ortho/meta-位置而发生亲核加成反应。4）PBDEs与BphA1能够发生有效亲合且对调节微生物降解性能具有重要意义，亲合作用与降解速率常数间呈现指数型正相关关系。5）成功发展了PBDEs好氧微生物降解的2D/3D-QSARs，构效关系表明PBDEs与BphA1之间的静电作用主导调控降解性能，而疏水作用也不容忽视。meta/para-位取代，有助于提高静电与疏水作用，从而提升降解速率。PBDEs好氧微生物降解性能预测与分子作用机制阐释，可为高效降解菌的选育与污染修复提供科学依据。

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