Excellence in Research in Spectroelectrochemical Measurements in Intact Microorganisms

询问固体表面和完整微生物之间发生的化学反应的常见手段通常以某种方式使用光来监测伴随这种反应的颜色或荧光变化。 固体和完整微生物的表面都使光偏转，从而干扰任何光学测量。 该项目将提供并验证一种新的方法，对固体和活微生物之间的反应进行光测量。 将使用一种带有观察室的新仪器，该仪器允许收集和测量所有光测量，而不是简单地丢失或忽略反应中固体或颗粒偏转的光。 该项目将重点关注5至10种微生物，它们与外部固体表面自然交换电子，作为其生活方式的一部分。 预期具有不同类型外部结构的微生物将使用不同策略和有色生物分子与外部固体表面交换电子。 该项目将提供一种新的手段来研究非侵入性生理条件下的生物能量转导，而不破坏活细胞环境的复杂性。 一个直接的结果将是，该项目将提供优化微生物燃料电池（新型电池）在阳极和阴极的性能特征所需的基本信息。 本科少数民族学生将在两个实验室的每一个为这个项目。 总体而言，至少有8名或更多的少数民族学生将接受培训。 该项目的长期目标是确定这些细菌中的电子传递生物分子和呼吸途径，这些细菌在强酸性条件下从可溶性和不溶性铁的有氧和厌氧呼吸中获得生长能量。 将同时进行光谱电化学测量和动力学研究，以确定选定的微生物在电解/观察池组合中呼吸时的耗氧速率。 为了实现这些目标，该项目将实施一种新的方法，该方法首次能够在生理条件下对活细胞中的原位电子转移反应进行直接分光光度测量，因为它们直接或间接地通过可溶性铁接受和捐赠电子到浸没在观察细胞中的电极。 这些研究将利用一种新型的积分腔吸收计，该吸收计具有创新的球形反射腔作为其样品观察室。 由于在腔壁处的多次反射，由于防止散射光逸出检测器，因此消除或最小化了由于样品浊度引起的散射损失。 这些组合的光谱和氧测量将在完整微生物在可逆循环伏安法电位扫描期间氧化和还原的同时进行。 待研究的微生物的属包括但不限于嗜酸硫杆菌属（Acidithiobacillus）、嗜酸嗜盐菌属（Leptocellulum）、嗜酸盐菌属（Acidihalobacillus）、硫杆菌属（Sulfobacillus）、铁浆菌属（Ferroplasma）和金属球菌属（Metallosphaera），它们代表含有呼吸铁的成员的6个门中的5个门。 先前的原位吸光度测量表明，这五种微生物中的每一种似乎表达一组不同的有色电子转移生物分子，以在铁上有氧呼吸。 因此，这些研究提供了机会，比较和对比5种不同的机制/途径，以实现相同的代谢目标，呼吸可溶性铁。这个奖项反映了NSF的法定使命，并已被认为是值得通过评估使用基金会的知识价值和更广泛的影响审查标准的支持。

Common means of interrogating chemical reactions that occur between solid surfaces and intact microorganisms typically use light in some manner to monitor either color or fluorescence changes that accompany such reactions. The surfaces of both the solids and the intact microorganisms deflect light and thereby interfere with any optical measurements. This project will provide and validate a new means to conduct light measurements on reactions between solids and live microorganisms. A new instrument with an observation chamber that permits all the light measurements to be harvested and measured will be used, rather than simply losing or ignoring the light that is deflected by the solids or particles in the reaction. This project will focus on 5 to 10 microorganisms that naturally exchange electrons with exterior solid surfaces as part of their lifestyle. It is anticipated that microorganisms with different types of exterior architectures will exchange electrons with exterior solid surfaces using different strategies and colored biomolecules. This project will provide a new means to study biological energy transduction under noninvasive physiological conditions without disrupting the complexity of the live cellular environment. One immediate outcome will be that this project will provide the basic information required to optimize the performance characteristics of microbial fuel cells (novel batteries) at both the anode and the cathode. Undergraduate minority students will be employed in each of 2 laboratories for this project. Overall, a minimum of 8 or more minority students will be trained. The long-term goal of this project is to define the electron transport biomolecules and respiratory pathways in those bacteria that obtain energy for growth from aerobic and anaerobic respiration on soluble and insoluble iron under strongly acidic conditions. Simultaneous spectroelectrochemical measurements and kinetic studies will be conducted to determine the rates of oxygen consumption as selected microorganisms respire in a combination electrolytic/observation cell. To accomplish these goals, this project will implement a new methodology that makes it possible, for the first time, to conduct direct spectrophotometric measurements of in situ electron transfer reactions in living cells under physiological conditions as they accept and donate electrons either directly or indirectly via soluble iron to electrodes immersed in the observation cell. These studies will utilize a novel integrating cavity absorption meter that has an innovative spherical reflecting cavity as its sample observation chamber. Due to multiple reflections at the cavity wall, scattering losses due to sample turbidity are eliminated or minimized because scattered light is prevented from escaping the detector. These combined spectral and oxygen measurements will be conducted while the intact microorganisms are oxidizing and reducing during reversible cyclic voltammetry potentiometric scans. The genera of microorganisms to be studied include, but are not limited to, Acidithiobacillus, Leptospirillum, Acidihalobacter, Sulfobacillus, Ferroplasma, and Metallosphaera, representing 5 of the 6 phyla that contain members that respire on iron. Prior in situ absorbance measurements demonstrated that each of these five microorganisms appeared to express a different set of colored electron transfer biomolecules to respire aerobically on iron. Thus, these studies provide the opportunity to compare and contrast 5 different mechanisms/pathways to accomplish the same metabolic goal of respiring on soluble iron.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.