微生物电还原重金属污染物的多重反向电子传递途径解析及调控研究

Heavy metal pollution is one of the biggest environmental problems that seriously threatened the human health worldwide. Heavy metal removal is a major challenge for wastewater treatment due to its high energy-consumption and low efficiency. Microbial electro-reduction is a promising new technology for pollutants treatment owing to its unique features of high efficiency and energy-generation. Reverse/inwards electron transfer is the newly explored microbial energy metabolism process, and underlies the microbial electro-reduction process. However, it is difficult (if not impossible) to rationally manipulate this new process as the detailed mechanism/pathway for reverse/inwards electron transfer is still unclear. . By using Shewanella oneidensis MR-1 as the model microorganism, Cr(VI) as the model heavy metal pollutant, we found that this bacterium could efficiently reduce Cr(VI) to Cr(III) using cathodic electrons as the sole electron source. Moreover, experimental results implied that there are more than one reverse electron transfer pathways played crucial roles in this Cr(VI) microbial electro-reduction process by S. oneidensis MR-1. Therefore, we will further investigate the effect of reverse electron transfer on the microbial electro-reduction of Cr(VI), and reveal the roles of reverse electron transfer in this bioelectro-reduction process. By using system biology tools (genome-wide transposon mutagenesis and comparative transcriptome), function verfication with heterogenesis expression, and bioinformatics approaches, the genes responsible for the reverese electron transfer will be elucidated, the relationship among these pathways and possible regulatory networks will be explored. More improtantly, a multi-pathway model for Cr(VI) induced reverse electron transfer will be proposed and validated with different methods (genetic and engineering) and at different levels (transcription, protein, physiology). Further, new rational designed highly efficient reverse electron transfer pathway(s) will be constructed and new manipulation strategy for efficient microbial electro-reduction of Cr(VI) will be proposed and validated to greatly improve the efficiency for Cr(VI) reduction. . This project will provide new insight into the molecular mechanism of microbial reverse electron transfer, develop new principle for rational manipulation of microbial reverse electron transfer, explore new strategies for efficient controlling of microbial electro-reduction process. These new knowledge and methods to be delivered will add new dimension to sustainable heavy metal pollution treatment.

重金属污染是重大环境问题，耗能、低效是其治理技术的关键瓶颈。反向电子传递是重要的微生物新过程，它驱动的微生物电还原是高效、产能的污染治理新技术，但反向电子传递途径尚不清楚，难以有效调控。我们前期以希瓦氏菌为模式菌，以首要重金属污染物Cr(VI)为典型污染物，研究发现有多重反向电子传递途径在重金属的微生物电还原过程起关键作用。据此，本项目将系统研究反向电子传递对Cr(VI)微生物电还原的作用及规律；采用全基因组突变体库、转录组和异源途径重建分析，筛选反向电子传递途径相关基因，解析各途径之间的相互关系及调控模式，构建并验证多重反向电子传递途径的结构和调控模型；理性构建高效新途径；在此基础上，探索微生物电还原的调控规律及高效策略，大幅提高Cr(VI)的微生物电还原效率。该研究将有助于揭示微生物反向电子传递机制、创新微生物电还原调控理论，为突破重金属污染治理的关键瓶颈提供新理论、新思路和新方法。

铬污染量大、面广、危害严重，是首要重金属污染之一。Cr(VI)是国际公认的三种致癌金属物质之一，是对人体危害最大的8种化学物质之一，已被列为优先控制污染物。我国工业废水的铬总排放量居五种有毒重金属污染物之首。传统生物还原法存在污泥量大、效率低、成本高等严重问题。因此，开发节能、高效的新型Cr(VI)还原技术是实现铬污染治理可持续发展的必由之路。本研究结合微生物电化学技术，开发了微生物电还原Cr(VI)技术，初步阐明了MtrCAB电子传递链是Cr(VI)生物电还原过程电子正反传递的主要途径，同时发现了MtrDEF和PsrABC硫还原酶、氢化酶HydA和HyaB及电子传递中介体在Cr(VI)生物电还原过程中的作用，建立了电子传递途径模型。建立了希瓦氏菌反向电子传递定量监测新方法，进一步定量阐明了电子传递链不同蛋白CymA, MtrA, MtrB, MtrC, OmcA对反向电子传递的调控规律；解析了核黄素对反向电子传递及Cr(VI)生物电还原调控的定量关系，发现了吩嗪化合物对反向电子传递的特殊调控作用及规律。进一步，研究发现细菌对Cr(VI)主要以胞内还原为主是导致Cr(VI)细胞毒性大、还原效率低的主要原因，提出了将Cr(VI)胞内还原转变为胞外生物电还原的新思路。在此基础上，提出了一系列希瓦氏菌电子传递及Cr(VI)生物电还原调控策略，使电子传递效率和污染物去除效率得到大幅提高；同时，提出了生物纳米杂合体系的新策略，将Cr(VI)胞内还原转变为胞外生物电还原，使Cr(VI)生物还原速率提高约8.4倍。基于以上研究，共发表SCI论文20篇，申请国家发明专利2项，培养硕士毕业生5人，2人获江苏省优秀硕士论文，联合培养外籍博士毕业生1人，圆满完成项目各项任务。