金属蛋白-纳米复合物用于构建高选择性传感分析系统

The world-wide heavy metal pollution issue has attracted extensive attention for the development of sensitive and rapid analysis/sensoring system. Metal sensing methodologies based on nanomaterials have been booming due to the advantages of simplicity and high sensitivity. The recognition of target metal species has long been a challenging task in metal sensing. Metalloregulatory protein/metal binding protein coded by metal resistance gene inside microbial cells can recognize their corresponding target metals with high selectivity and affinity. Meanwhile, the outstanding optical properties of nanomaterials further endow them to be able to detect the targets with good sensitivity. The combination of the two features will definitely open a new avenue for highly sensitive recognition and sensing of heavy metals and their species. .The present project will focus on the following issues: to express metal binding protein/peptide and metalloregulatory protein inside microbial cells by gene engineering technique; to investigate the interactions between metal species and proteins in order to elucidate the possible chemical/physical changes; to combine metal proteins with nanomaterials (graphene, carbon nanostructutres, noble metal nanoparticles, etc.), and to construct the metal protein-nano composites for the development of highly selective optical sensing system for heavy metals based on the mechanism of fluorescence quenching, fluorescence resonance energy transfer (FRET) or surface enhanced raman scattering (SERS).

重金属污染问题的日益凸显加速了重金属灵敏快速分析的需求。基于纳米材料的传感方法以其简便、灵敏、快速等优势得到了迅速发展。重金属传感的难点在于目标金属的识别。微生物体内重金属拮抗基因编码的金属调节蛋白/金属结合蛋白对目标金属离子具有高特异性的识别能力。将特异性识别金属离子的金属蛋白与具有优异光谱性能的纳米材料结合，有望为重金属的高选择性识别传感开辟新的途径。.本项目旨在运用分子生物学手段，在微生物细胞体内表达合适的金属结合蛋白/多肽及金属调节蛋白，并将其纯化，研究这些蛋白/多肽与重金属形态的作用机理以及作用过程中物理化学性质的改变；与石墨烯、碳纳米结构、贵金属纳米粒子等纳米材料结合，构建金属蛋白-纳米复合体系，研究体系对重金属形态的选择性及特异性响应，并基于荧光猝灭、荧光共振能量转移或表面增强拉曼等机理，建立新型高选择性重金属离子的光学传感分析系统。

在自然科学基金青年基金“金属蛋白-纳米复合物用于高选择性传感分析系统的构建”（21405010）资助下，我们通过噬菌体多肽文库技术，筛选获得了多种对目标金属具有特异性结合能力的噬菌体及对应多肽，并基于此建立了一系列高选择性重金属比色传感方法。主要研究内容包括：（1）亲汞噬菌体原位合成纳米金及其在汞传感分析中的应用研究。以表面展示汞结合多肽的M13噬菌体为生物模板，原位合成金纳米颗粒，并以噬菌体-纳米金交联复合体系为传感平台，建立起汞离子的选择性捕获、还原、传感分析新策略；（2）砷结合肽-纳米金传感体系选择性检测As(III)。利用As(III)与金纳米颗粒竞争性结合砷结合多肽过程中产生的表面等离子体共振光谱变化，建立起灵敏、快速的As(III)比色传感方法；（3）铬结合七肽的噬菌体文库淘选及其应用研究。通过噬菌体文库技术淘选获得铬结合七肽，并以表面展示铬结合七肽的M13噬菌体为固相萃取材料，建立起Cr(III)的选择性分离富集及无机铬形态分析方法。项目负责人作为第一作者和通讯作者发表基金标注SCI收录论文11篇，其中影响因子大于7的论文3篇，包括1篇Nanoscale、1篇ACS Applied Materials & Interfaces、1篇TrAC Trends in Analytical Chemistry。项目在研究成果与人才培养方面已达到预期目标。