包气带微生物对多环芳烃的跨膜运输及膜蛋白传输方式的研究

According to the complex relation among aeration zone-microorganism-polycyclic aromatic hydrocarbons (PAHs) system, the key links and rate-control steps in the biodegradation process of PAHs were not clear yet. Two species of PAHs-degrading bacteria (Pseudomonas sp.DG17 and Rhodococcus sp. BAP-1) that we isolated from oil-contaminated soils in aeration zone before were used as the target cells. Processes of the trans-membrane transport of PAHs (phenanthrene, pyrene, fluoranthene, benzo[a]pyrene, benzo[b]fluoranthene) outside the membrane, across the periplasm and inside the cell were studied by using the isotope tracing technique. Meanwhile, the specie, function, structure and transmission mode of the membrane protein were analyzed via NMR and NSLS methods, also the binding process and binding site for PAHs and membrane protein were investigated. Based on the studies, the key links and rate-control steps in the biodegradation process of PAHs by using the bacteria that isolated from the aeration zone were determined; the relationships between the environmental conditions of aeration zone and the improvements of the biodegradation rate of PAHs were explored. The achievements will be used to provide the theoretical support and practical guidance in bioremediation of PAHs-contaminated soils in aeration zone.

针对包气带- - 微生物- - 多环芳烃降解体系作用关系复杂，多环芳烃在降解过程中的关键环节和控速步骤尚不明确。本研究以课题组在前期研究中筛选出的高效多环芳烃降解菌：假单胞菌DG17和红球菌BAP-1为研究靶细胞，利用同位素示踪技术研究多环芳烃（菲、芘、荧蒽、苯并[a]芘、苯并[b]荧蒽）在微生物细胞外、细胞膜周及细胞内的分配及跨膜运输过程；在此基础上，通过核磁共振和同步辐射光源法对微生物跨膜蛋白的种类、作用、结构和传输方式，以及多环芳烃与膜蛋白的结合过程进行分析。确定包气带微生物跨膜运输多环芳烃的关键环节和控制因子，探索不同包气带环境条件与提高微生物对多环芳烃降解效率之间的关系，为利用微生物技术修复包气带多环芳烃类污染物提供重要的理论依据和实践指导。

本项目以课题组前期研究中筛选出的高效多环芳烃降解菌：红球菌BAP-1和芽孢杆菌DG24为研究靶细胞，利用同位素示踪技术，完成了红球菌BAP-1对荧蒽以及芽孢杆菌DG24对菲的跨膜运输方式及跨膜运输动力学过程的研究，分析了不同包气带环境的变化与提高微生物跨膜运输多环芳烃类物质效率之间的关系，在此基础上确定了多环芳烃类污染物微生物跨膜运输过程的关键环节和限速步骤。探索了利用稳定同位素标记结合Nano-SIMS技术分析葡萄糖在微生物BDG-3体内代谢过程的实验条件。同时，建立了微生物在多环芳烃诱导下分泌细胞膜蛋白的提取定量与分析鉴定方法体系，并利用iTRAQ技术结合LC-MS/MS对荧蒽诱导下红球菌BAP-1细胞膜蛋白的差异表达进行了生物信息分析。为利用微生物技术及分子生物学技术修复包气带多环芳烃类污染物提供重要的理论依据和实践指导。