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

描述(由申请人提供)： 土壤和地下水的铬(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地的第三种常见污染物，也是环境中发现的第二种最常见的无机污染物。利用异化金属离子还原细菌(DMRB)固有代谢能力的原位和非原位生物修复过程仍然是还原固定化或环境污染物解毒的有效、潜在的经济有效的方法。例如，微生物催化将铬(VI)还原为难溶、生物有效性较低的铬(III)，是一种很有前途的修复策略，适用于受铬(VI)污染的地下土壤和地下水环境。希瓦氏菌属是少数几类受到深入研究的微生物之一，因为它们具有广泛的生态分布、不同的呼吸能力和与环境相关的特性。尽管在阐明希瓦氏菌与铬酸盐转化有关的生物学方面取得了一些进展，但关于特定的铬酸盐还原机制的基本问题仍然不清楚。这种信息差距包括(I)专用铬酸盐还原酶(S)的同一性，(Ii)铬酸盐转化的细胞定位(例如，远端附属物、细胞外表面、周质、细胞膜、胞浆)，以及(Iii)微生物群体具有最大特定铬酸盐还原速率的环境参数。预测和评估生物修复性能的问题因缺乏使细菌还原金属能力的分子基础、调节机制和生物化学的基本知识而变得更加复杂。我们建议设计纳米尺度的方法，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表面增强共振拉曼散射探测的增强剂，以生成铬酸盐还原位置的化学地图，以及以精细的分子和单个生物的细节监测还原动力学。这项研究的目标是：1)评估金纳米颗粒的组成、几何形状和功能对细胞活力、生长和微生物还原铬酸盐的有效性的影响；2)使用功能化的金纳米结构和使用细胞内生长的金纳米岛通过拉曼化学成像跟踪单细胞分辨率下铬酸盐转化的定位；以及3)评估与生物修复相关的环境因素对铬酸盐运输、定位和还原速率的影响。与共聚焦拉曼化学成像相结合的被动和主动纳米探针的开发，将成为能够以单细胞分辨率动态监测细胞内事件和划分金属还原部位的平台的重要一步。从这项新颖的研究中获得的知识将有助于制定以科学为基础的战略，以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.