Structure/Function Correlations Over Binuclear Non-Heme Iron and Related Enzymes

生物体利用双核非血红素铁酶促进O2的广泛化学反应。例如，人类需要核糖核苷酸还原酶来生成DNA构建块，而细菌种类使用可溶性甲烷单加氧酶将甲烷（一种温室气体）转化为甲醇（一种可用的燃料）。这些酶已成为开发抗癌药物和生物燃料催化剂的目标。在分子水平上理解它们的化学性质需要阐明催化反应中铁活性核的结构变化；然而，其中许多是用传统的光谱技术无法获得的。因此，这个项目包括开发新的光谱方法，利用强磁场、圆偏振光和第三代同步辐射。实验数据结合量子力学计算，确定六种酶的反应机理和相关模型。从这个项目中获得的分子水平的洞察力将促进药物发现和生物技术。所开发的方法对许多科学领域产生了影响，并通过许多国家和国际合作提供，这些合作现在正在扩大到仅限本科生的美国机构。该项目还包括多种推广工作，包括高中教师/学生参与我们实验室的夏季研究和其他K-12项目。科学计划培养了一大批非常成功的科学家，并编辑了重要的科学出版物。双核非血红素酶形成扩展类，激活O2进行h原子提取、去饱和、羟基化和亲电攻击，并将O2还原为H2O，将NO还原为N2O。PI一直在开发光谱方法来探测它们的O2活化还原态（变温/变场磁圆二色性，VTVH MCD）和它们的氧中间体（核共振振动光谱，NRVS）。现在的重点是：1)了解1e-还原（肌醇加氧酶，激活O2作为结合超氧化物的原型）激活O2的机制；2)开发/使用NRVS/MCD来定义过氧化物- 2feiii中间体的性质和活化；3)将Aim 2扩展到可溶性甲烷单加氧酶的高价中间体Q和核糖核苷酸还原酶的高价中间体X；4)明确黄素二铁蛋白及其他相关蛋白中O2和NO还原的机制；5)继续开发/使用VTVH MCD和其他光谱来了解这些酶的新类别的结构/功能相关性。这些对广泛的双核铁酶的研究建立了多中心酶用于O2活化和指导催化的一般原理。

Organisms utilize binuclear non-heme iron enzymes to facilitate a wide range of chemical reactions using O2. As examples, humans need ribonucleotide reductase to generate DNA building blocks, and classes of bacteria use soluble methane monooxygenase to convert methane (a greenhouse gas) to methanol (a usable fuel). These enzymes have been targeted for the development of anti-cancer drugs and biofuel catalysts. Understanding their chemistry at a molecular level requires elucidating structural changes of the iron active core over the catalytic reaction; however, many of these are not accessible with traditional spectroscopic techniques. Thus, this project includes the development of new spectroscopic methodologies that utilize intense magnetic fields, circularly polarized light, and third-generation synchrotron radiation. Experimental data combined with quantum mechanical calculations will define reaction mechanisms of six enzymes and related models. The molecular level insight gained from this project will promote drug discovery and biotechnology. The methods developed have impact on many areas of science and are made available through numerous national and international collaborations, which are now expanding to undergrad-only U.S. institutions. This project also includes multiple outreach efforts, including participation of high school teachers/students in summer research in our labs and other K-12 programs. The PI has trained a large number of highly successful scientists and edits major scientific publications.The binuclear non-heme enzymes form expanding classes that activate O2 for H-atom abstraction, desaturation, hydroxylation, and electrophilic attack, and reduce O2 to H2O and NO to N2O. The PI has been developing spectroscopic methodologies to probe their O2 activating reduced states (variable-temperature/variable-field magnetic circular dichroism, VTVH MCD,) and their oxygen intermediates (nuclear resonance vibrational spectroscopy, NRVS). The focus is now on: 1) understanding the mechanism of O2 activation by 1e- reduction (myoinositol oxygenase, the prototype that activates O2 as bound superoxide); 2) developing/using NRVS/MCD to define the nature and activation of peroxo-2FeIII intermediates; 3) extending Aim 2) to the high-valent intermediates Q in soluble methane monooxygenase and X in ribonucleotide reductase; 4) defining the mechanisms of O2 and NO reduction in flavin diiron proteins and other related classes; 5) continuing to develop/use VTVH MCD and other spectroscopies to understand structure/function correlations in new classes of these enzymes. These studies over a wide range of binuclear iron enzymes establish general principles utilized by multicenter enzymes for O2 activation and in directing catalysis.