Chemical Mapping of Chromate Uptake, Localization, and Reduction in Remediating B

描述（由申请人提供）： 土壤和地下水的Cr（VI）污染是一个世界性的重大问题。在美国，铬酸盐是危险废物场的第三大常见污染物，也是环境中第二大常见无机污染物。利用异化金属离子还原菌（DMRB）的内在代谢能力的原位和异位生物修复过程仍然是有效的，潜在的成本效益的方法来还原固定或解毒的环境污染物。例如，微生物催化将Cr（VI）还原为微溶的、生物可利用性较低的Cr（III）是Cr（VI）污染的地下土壤和地下水环境的一种有前途的修复策略。希瓦氏菌属代表了少数几组微生物之一，因为它们广泛的生态分布，多样的呼吸能力和环境相关性而受到深入研究。尽管在阐明希瓦氏菌生物学方面取得了一些进展，因为它与铬酸盐转化有关，但有关特定铬酸盐还原机制的基本问题仍不清楚。这种信息缺口包括（i）专用铬酸盐还原酶的身份，（ii）铬酸盐转化的细胞定位（例如，远端附属物，外细胞表面，周质，细胞质膜，胞质溶胶），和（iii）环境参数下，微生物种群具有最大的比铬酸盐还原率。预测和评估生物修复性能的问题是由于缺乏分子基础、调控机制和生物化学方面的基本知识而加剧的，这些知识使细菌的金属还原能力成为可能。我们建议设计纳米级方法，包括铬酸盐标记的纳米颗粒和细胞内生长的金纳米岛作为增强剂的表面增强共振拉曼散射探测生成铬酸盐还原位点的化学图谱，以及监测精致的分子和单一生物体细节的还原动力学。本研究的目的是：1）评估金纳米颗粒的组成，几何形状和功能对细胞活力，生长和微生物铬酸盐还原效率的影响，使用S。oneidensis作为一个模型系统; 2）通过拉曼化学成像，使用功能化的金纳米结构以及使用细胞内生长的金纳米岛，以单细胞分辨率跟踪铬酸盐转化的定位;以及3）评估生物修复相关的环境因素对铬酸盐转运、定位和还原速率的影响。与共焦拉曼化学成像相结合的被动和主动纳米探针的开发将构成一个重要的步骤，使一个平台，用于动态监测细胞内的事件和划分的金属还原位点在单细胞分辨率。从这项新的研究中获得的知识将有助于发展科学的战略，提高生物修复的效果。

DESCRIPTION (provided by applicant): Cr(VI) contamination of soil and groundwater is a significant problem worldwide. In the United States, chromate is the third most common contaminant of hazardous waste sites and the second most common inorganic contaminant found in the environment. In situ and ex situ bioremediation processes that exploit the intrinsic metabolic capabilities of dissimilatory metal ion-reducing bacteria (DMRB) remain potent, potentially cost-effective approaches to the reductive immobilization or detoxification of environmental contaminants. The microbial catalysis of Cr(VI) reduction to sparingly soluble, less bioavailable Cr(III), for example, is a promising remediation strategy for Cr(VI)-contaminated subsurface soil and groundwater environments. The genus Shewanella represents one of the few groups of microorganisms that have received intensive investigation because of their wide ecological distribution, diverse respiratory capacities, and environmental relevance. Despite several advances made in elucidating Shewanella biology as it relates to chromate transformation, fundamental questions about the specific chromate reduction mechanism remain unclear. This information gap includes (i) the identity of dedicated chromate reductase(s), (ii) the cellular localization of chromate transformation (e.g., distal appendages, outer cell surface, periplasm, cytoplasmic membrane, cytosol), and (iii) the environmental parameters under which microbial populations have the greatest specific chromate reduction rates. The problem in predicting and assessing bioremediation performance is compounded by the lack of fundamental knowledge of the molecular basis, regulatory mechanisms, and biochemistry enabling bacterial metal-reducing capabilities. We propose to engineer nanoscale methodologies, comprising of chromate-tagged nanoparticles and intracellularly grown gold nanoislands to function as enhancers for Surface Enhanced resonance Raman scattering probing to generate chemical maps of chromate reduction sites as well as to monitor the reduction dynamics in exquisite molecular and single-organism detail. Objectives of this study are to 1) assess the impact of gold nanoparticle composition, geometry, and functionality on cell viability, growth, and efficacy of microbial chromate reduction using S. oneidensis as a model system; 2) track the localization of chromate transformation at single-cell resolution using functionalized gold nanostructures as well as using intracellularly grown gold nanoislands by Raman chemical imaging; and 3) evaluate the influence of bioremediation-relevant environmental factors on chromate transport, localization, and reduction rates. The development of passive and active nanoprobes in conjunction with confocal Raman chemical imaging will constitute a significant step in enabling a platform for dynamic monitoring of intracellular events and compartmentalization of metal reduction sites at single-cell resolution. The knowledge gained from this novel study will contribute to the development of scientifically grounded strategies for improving bioremediation efficacy.