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

生物体利用双核非血红素铁酶来促进利用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构件，而各类细菌使用可溶的甲烷单加氧酶将甲烷(一种温室气体)转化为甲醇(一种可用燃料)。这些酶已被用于开发抗癌药物和生物燃料催化剂。在分子水平上了解它们的化学需要阐明铁活性核心在催化反应中的结构变化；然而，其中许多变化是传统光谱技术无法获得的。因此，该项目包括开发利用强磁场、圆偏振光和第三代同步辐射的新光谱方法。实验数据结合量子力学计算将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。开发的方法对许多科学领域都有影响，并通过许多国家和国际合作提供，这些合作现在正扩展到仅限本科生的美国机构。该项目还包括多项外展工作，包括高中教师/学生参与我们实验室的暑期研究和其他K-12项目。PI培养了一大批非常成功的科学家和编辑了主要的科学出版物，双核非血红素酶形成了扩展的类别，激活O2进行H原子的提取、脱饱和、羟化和亲电攻击，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索它们的氧活化还原态(变温/变场磁圆二色谱，VTVH MCD)和它们的氧中间体(核共振振动光谱，NRVS)。目前的重点是：1)了解1e-还原激活O2的机制(肌醇加氧酶，激活O2作为结合的超氧化物的原型)；2)开发/使用NRVS/MCD来定义Peroxo-2FeIII中间体的性质和激活；3)将目标2)扩展到可溶性甲烷单加氧酶中的高价中间体Q和核糖核苷酸还原酶中的X；4)定义黄素二铁蛋白和其他相关类别中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中的结构/功能相关性。这些对广泛的双核铁酶的研究确立了多中心酶用来激活O2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.