CAREER: Decoding Iron-Catalyzed Oxidation Mechanisms in Biology via Time-Resolved X-ray Approaches

Researchers from Emory University are developing and applying new approaches to understand how iron-containing enzymes harness highly reactive forms of oxygen to carry out chemical reactions essential for biological processes ranging from natural product biosynthesis to environmental detoxification. Studying critical intermediate states of these reactions is challenging because they often persist for very short timescales, making them difficult to observe directly and leaving unanswered fundamental questions about how oxygen-activating enzymes control reactivity. The research team will use light to trigger these reactions under controlled conditions combined with advanced X-ray methods to generate molecular movies of enzyme catalysis that correlate structural dynamics with changes at the metal center. Their discoveries could establish a broadly applicable framework for understanding biological oxidation chemistry, with implications for biotechnology, sustainable catalysis, human health, and environmental remediation. The project will also support public outreach through partnerships with Science ATL and curricular innovations that strengthen undergraduate physical chemistry education, while providing interdisciplinary research training opportunities. This award will investigate the mechanisms of mononuclear non-heme iron enzymes by directly characterizing ferryl and ‘bridged’ peroxo intermediates. Photoinitiation methods will be developed and applied in combination with serial crystallography and X-ray spectroscopy to study reactive intermediates in representative Fe(II) and 2-oxoglutarate-dependent oxygenases and aromatic amino acid hydroxylases. These studies will determine whether O-O bond cleavage and ferryl formation follow a conserved geometric trajectory or reflect distinct structural adaptations tuned to respective catalytic contexts. They will also establish how oxidation state and spin state changes are coupled to protein conformational dynamics. The resulting structural and electronic information will advance understanding of substrate activation and reaction specificity across these enzyme families. In parallel, the experimental platform advanced through this work will provide a modular strategy that is readily adaptable for time-resolved studies of diverse redox-active metalloenzymes, expanding the mechanistic toolkit available for structure-function investigations of complex biological oxidation reactions. 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.