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

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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.