Collaborative Research: Biochemical and Spectroscopic Studies of Mononuclear Iron Enzyme Catalyzed C-X Bond Forming and Cleaving Reactions using O2 as the only Co-Substrate

North Carolina State University, Raleigh NC

Investigators

Abstract

With the support of the Chemistry of Life Processes (CLP) program in the Division of Chemistry, Professors Yisong Guo of Carnegie Mellon University and Wei-chen Chang of North Carolina State University will use biochemical and spectroscopic tools to investigate a group of four enzymes, which utilize a single iron and molecular oxygen (O2) to catalyze diverse while synthetically challenging chemical reactions to make valuable small molecules. These reactions have the potential to: (1) convert precursors to the building blocks of antibiotics or potential antimalarial agents; and 2) generate molecules that exhibit physiological functions in vision, light harvesting, and pathogen virulence, and some of them are linked to cardiovascular disease. The goal of this research program is to elucidate the fundamental knowledge on how these enzymes utilize iron and O2 to achieve diverse reaction outcomes. The obtained results will set the foundation for developing biocatalytic platforms to access these important chemical transformations to benefit modern society. The research goals will be integrated into educational activities to disseminate cutting-edge scientific results and research skills to graduate level, undergraduate level, and K-12 level students via course and outreach program development. Over the past decades, the mechanistic understanding of co-substrate-dependent (e.g., 2-oxoglutarate-dependent and pterin-dependent) non-heme mononuclear iron-containing (NHM-Fe) enzymes has been greatly improved. One key factor empowering these progresses is the identification and characterization of the common reactive intermediate (e.g., the S = 2 oxoiron(IV) intermediate) in co-substrate-dependent enzymes. However, key reactive intermediate(s) for co-substrate-independent NHM-Fe enzymes have not been experimentally verified, and it is still unclear regarding how O2 activation takes place without the involvement of a co-substrate. In this research program, the Guo and the Chang groups will work as a team to reveal detailed reaction mechanisms of four newly discovered co-substrate-independent NHM-Fe enzymes via the identification and characterization of iron-based reactive intermediates and reaction outcome analysis. These enzymes exhibit diverse catalytic activities including carbon-carbon bond formation and breakdown, and carbon-nitrogen bond formation. To achieve the research goals, an integrated research approach, consisting of substrate probe design and synthesis, transient enzyme kinetics, electron paramagnetic resonance (EPR), Mössbauer, and X-ray absorption spectroscopy (XAS) will be used. The use of this suite of tools could lead to a comprehensive understanding of the factors controlling O2 activation and reaction outcomes at a molecular level, which would fill the crucial knowledge gap in the understanding of NHM-Fe enzyme catalysis beyond the co-substrate dependent enzymes. 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.