Bacterial Exudates: Characteristics and Role in Selenium Reduction

The goals of this research are to further our understanding of interactions between soluble bacterial exudates and selenium, and to determine how these interactions may affect the environmental behavior of Se.

In Chapter 2, I discuss a study that tests whether soluble bacterial exudates can reduce aqueous Se(IV). Exudate solutions were isolated from three different model species of bacteria and tested for reductive activity to added Se(IV). The results show that some bacterial exudates reduce Se(IV), but that those from Shewanella oneidensis can not unless the cells are grown in the presence of aqueous Se(IV). The results also demonstrate that Se(IV) reduction by each of the exudate solutions involves exudate-hosted sulfhydryl binding.

Chapter 3 describes potentiometric titration experiments that were used to measure the concentrations of proton-active sites on the exudate molecules produced by several bacterial species as a function of soak time, and the chapter also reports the first measurements of sulfhydryl site concentrations on exudate molecules. The total binding site concentrations of the exudates are primarily controlled by DOC concentration, unlike sulfhydryl site concentrations, suggesting that production of sulfhydryl-site bearing exudates is primarily an active process.

The research reported in Chapter 4 uses comparative proteomics to identify candidate proteins that are responsible for the inducible Se(IV) reductase activity of soluble S. oneidensis exudates. Combining the control of S. oneidensis exudates’ inducible activity with knowledge of required characteristics for Se(IV) reduction allows us to narrow the list to six candidate proteins for further investigation.

The research in this dissertation demonstrates for the first time the ability of soluble bacterial exudates to reduce Se(IV) and suggests that Se(IV) binding onto sulfhydryl sites controls the reduction reaction. The research characterizes the total concentration of binding sites and the concentration of sulfhydryl sites specifically on exudate molecules, and it identifies six candidate proteins that are responsible for the observed Se(IV) reduction. Overall, the research presented here improves our understanding of Se(IV)-exudate reactions, and could be useful in refining environmental models of selenium behavior and developing bioremediation methods for selenium contamination.

本研究旨在深化我们对可溶性细菌分泌物与硒之间相互作用的认知，并探究这些相互作用可能对硒的环境行为产生的影响。在第二章中，我讨论了一项研究，旨在检验可溶性细菌分泌物是否能够降低水溶液中的硒(IV)浓度。从三种不同的细菌模型物种中分离出的分泌液被用于测试其相对于添加的硒(IV)的还原活性。结果显示，某些细菌分泌物能够还原硒(IV)，但来自希瓦氏菌属的分泌物则不能，除非在含有水溶液硒(IV)的环境中培养细胞。此外，结果还表明，每种分泌液对硒(IV)的还原过程均涉及分泌物中的硫醇基结合。第三章描述了电位滴定实验，用于测量多种细菌物种产生的分泌物分子上质子活性位点的浓度，并随浸泡时间的函数变化，该章节还报告了首次对分泌物分子上硫醇位点浓度的测量。分泌物的总结合位点浓度主要受溶解性有机碳（DOC）浓度控制，而硫醇位点浓度则不然，这表明带有硫醇位点的分泌物生产主要是一个主动过程。第四章中报道的研究利用比较蛋白质组学方法，以确定负责可溶性希瓦氏菌属分泌物诱导硒(IV)还原酶活性的候选蛋白质。结合对希瓦氏菌属分泌物诱导活性的控制以及对硒(IV)还原所需特征的了解，我们能够将候选蛋白质列表缩小至六个，以进行进一步的研究。本论文所报道的研究首次证明了可溶性细菌分泌物能够还原硒(IV)，并表明硒(IV)结合到硫醇位点上控制了还原反应。研究对结合位点的总浓度以及分泌物分子上硫醇位点的浓度进行了表征，并确定了六个负责观察到的硒(IV)还原的候选蛋白质。总体而言，本研究加深了我们对硒(IV)-分泌物反应的理解，并可能有助于改进硒的环境行为模型，以及开发硒污染的生物修复方法。