Developing Electroanalytical Methods for Enzymology Applications

在化学部(CHE)的化学测量与成像(CMI)项目和CBET部门的生物传感器项目的支持下，犹他大学的Shelley Minteer教授正在开发新的电分析工具，用于研究酶，生物催化剂在生命过程的化学中发挥关键作用，但在生物技术中也非常有用。生物化学家和分子生物学家对酶结构如何控制功能产生了浓厚的兴趣。此外，酶还启发了选择性催化剂的设计，用于挑战转化，如将二氧化碳还原为甲醇，将氮(N2)还原为氨。在过去的几十年里，这些属性推动了广泛的酶学研究，促使人们有兴趣开发更好的分析工具来研究酶的功能。Minteer小组正在开发这样的工具，并通过向大学生和高中生提供教育机会来扩大其影响。例如，Minteer教授正在提供独特的高中生研究实习机会，以激发学生对STEM(科学、技术、工程和数学)学科的兴趣。为了开发新的酶化学电分析方法，关键是能够将酶固定在电极表面，使它们能够与电极发生直接的电子转移，而不需要外源氧化还原介质。Minteer团队正在利用两种策略实现直接电子转移：1)将非天然氨基酸作为定向标签，将蛋白质连接到电极表面，从而确保活性部位位于电极表面的隧道距离内；2)通过结合金属结合的多肽序列，促进蛋白质内金属纳米颗粒的原位形成，将酶“连线”到电极表面。任何一种方法修饰的生物电极都将被用来研究单酶和使用新的恒流滴定方法的酶级联，目的是更好地理解电子注入蛋白质活性部位(即固氮酶)的速度如何控制形成的产物(氨和氢)的选择性。此外，旋转环-圆盘电分析化学方法正在开发中，以研究底物通道和定量中间体从酶级联的超复合体泄漏到主体溶液中。该奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

With support from the Chemical Measurement & Imaging (CMI) program in the Division of Chemistry (CHE) and the Biosensors program in the CBET Division, Professor Shelley Minteer at the University of Utah is developing new electroanalytical tools for investigating enzymes, biological catalysts that play critical roles in the chemistry of life processes but that are also very useful in biotechnology. Biochemists and molecular biologists have keen interest in how enzyme structure controls function. In addition, enzymes have inspired the design of selective catalysts for challenging transformations such as reduction of carbon dioxide to methanol and nitrogen (N2) to ammonia. These attributes have motivated extensive enzymology research over the last couple of decades, prompting interest in developing better analytical tools for investigating enzyme function. The Minteer group is both developing such tools and extending their impact through educational opportunities offered to both college and high school students. For example, Professor Minteer is providing unique high school student research internships to inspire student interest in STEM (science, technology, engineering and mathematics) disciplines. In order to develop new electroanalytical methods for enzymology, it is critical to be able to immobilize enzymes on electrode surfaces in a manner whereby they can undergo direct electron transfer with the electrode without the need for exogenous redox mediators. The Minteer group is utilizing two strategies to enable direct electron transfer: 1) incorporation of non-natural amino acids as orienting tags for conjugation of proteins to electrode surfaces, thereby ensuring that the active site is within tunneling distance of the electrode surface; and 2) "wiring" enzymes to electrode surfaces by incorporating metal-binding peptide sequences that promote in-situ formation of metal nanoparticles within the protein. Bioelectrodes modified by either approach will be used to study both single enzymes and enzyme cascades using new galvanostatic titration methods, with an aim of improved understanding of how the rate of electron injection into the active site of a protein (i.e. nitrogenase) controls the selectivity of the products formed (ammonia versus hydrogen). Additionally, rotating ring-disk electroanalytical chemistry methods are being developed to study substrate channeling and to quantitate the leaking of intermediates to the bulk solution from the supercomplex of an enzyme cascade.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.