新型生物还原法制备石墨烯过程的微生物群体感应调控及机制研究

Graphene is the strongest and thinnest nano-material which shows tremendous applications in various areas. However, serious environment pollution resulted from graphene synthesis process (chemical reduction of graphene oxide (GO), the most promising scalable strategy) hindered its commercial applications and becomes the bottleneck for graphene industry. Bioreduction of GO by bacteria holds great promise for green synthesis of graphene and removal of the toxic precursor (GO). Thus, understanding the microbial regulatory mechanism on GO bioreduction process is very important and crucial for advanced graphene industry. We previously isolated a Pseudomonas aeruginosa strain capable of GO reduction, and found that Rhl quorum sensing （QS）genetic system had positive regulation on the GO bioreduction process. Therefore, we intend to systematically investigate the effect of QS on the kinetics of GO bioreduction, and reveal the roles of Rhl quorum sensing in this bioreduction process. The relationship between the QS regulated physiological/metabolic behaviors and the GO reduction capability will be established, and the roles of QS regulated physiological/metabolic charaters in GO bioreduction process will be proposed and verified. By using system biology (genome-wide transposon mutagenesis and comparative transcriptome) and bioinformatics approaches, the gene targets and regulatory network for QS regulation on GO bioreduction will be revealed. By validating this regulatory network with different methods (genetic and engineering) and at different levels (transcription, protein, physiology), a comprehensive molecular mechanism of QS regulation on GO bioreduction process will be achieved. Further, new rational strategy for high efficient GO bioreduction, which based on the QS regulatory mechanism, will be proposed and carried out. This project will provide new understanding on the roles of QS in GO bioreduction, and will get novel insight on molecular mechanism of microbial regulation on GO bioreduction, which should be valuable fundamental information for rational design of high efficient strategy or synthetic super bug to approach green chemistry of graphene industry.

石墨烯是目前强度最大、厚度最小、用途广泛的纳米材料，但其规模化制备过程(氧化石墨烯(GO)化学还原)的高污染、高能耗已成为石墨烯工业发展的关键瓶颈。GO生物还原是推进石墨烯工业节能清洁生产的国际前沿技术，但其微生物调控机制尚不清楚，难以实现人工控制。我们前期已筛选到有GO还原能力的铜绿假单胞菌，发现其群体感应调控系统能够显著加速GO生物还原过程。据此，本项目将系统研究群体感应对GO生物还原的调控作用及规律；阐明群体感应调控GO还原的生理基础；采用全基因组突变体库和转录组分析，筛选群体感应调控GO还原的基因靶点；构建并验证群体感应调控GO还原的分子机制模型。在此基础上，提出人工理性调控策略，大幅提高GO生物还原效率。该研究将有助于建立GO生物还原过程的微生物调控理论、创新GO生物还原调控技术，为突破GO还原法制备石墨烯的关键瓶颈和促进石墨烯工业的可持续发展提供新思路和新方法。

石墨烯是目前强度最大、厚度最小、用途广泛的纳米材料，但其规模化制备过程(氧化石墨烯(GO)化学还原)的高污染、高能耗已成为石墨烯工业发展的关键瓶颈。GO生物还原是推进石墨烯工业节能清洁生产的国际前沿技术，但其微生物调控机制尚不清楚，难以实现人工控制。我们在前期发现有GO还原能力的铜绿假单胞菌的基础上，初步阐明了Rhl群体感应主要通过调控微生物电子传递过程来加速GO生物还原的调控机理，明确了吩嗪化合物PYO合成量与GO生物还原的相关性；建立了96孔板快速筛选GO生物还原能力改变的菌株的高通量方法，基于此方法通过全基因组突变体库筛选获得Rhl群体感应系统调控GO生物还原的靶标，从分子水平上揭示了吩嗪化合物合成基因簇（尤其是phzM基因）和鼠李糖脂合成酶(rhlA基因)是Rhl群体感应调控GO生物还原的主要靶标；明确了rhlA主要通过合成鼠李糖脂，提高吩嗪化合物PYO分泌与合成来促进GO生物还原的机制，并建立了Rhl调控GO生物还原的机理模型；据此，开发了rhlI/rhlR高表达、phzM高表达、槐糖脂添加、rhlA高表达等四种GO生物还原强化新策略，使GO生物还原速率大幅提高，GO污染生物修复速率提高4倍左右，石墨烯制备速率提高2.5倍左右，完成了项目预期研究目标。同时，在完成既定目标的基础上，基于GO生物还原机制，开发了GO生物还原过程在生物电化学体系的新应用。本研究共发表第一/通讯作者SCI论文10篇，其中包括IF>10的Angew. Chem. Int. Ed.1篇，IF>7的Biosens. Bioelectron. 1篇，IF=4.9的Bioresource Technol.3篇。撰写英文专著章节1章，申请发明专利3项，参加国内外学术会议交流报告10余次，得到了国际同行的高度评价。该研究将有助于揭示GO生物还原过程机制、创新了GO生物还原调控技术，为突破GO还原法制备石墨烯的关键瓶颈和促进石墨烯工业的可持续发展提供了新思路和新方法。