Elucidating mechanisms of biological hydrogen conversion through model metalloenzymes

With the support of the Chemistry of Life Processes Program in the Chemistry Division, Professor Hannah Shafaat of The Ohio State University will investigate the factors that govern the activity of nickel-containing enzymes. Reductive nickel enzymes are critical in the metabolic processes of diverse microorganisms and perform valuable reactions such as hydrogen production, carbon dioxide reduction, and methane oxidation. These efficient, complex enzymes operate with high rates and full reversibility but have yet to be accurately reproduced in synthetic systems, leaving many questions unanswered. To better understand native nickel enzymes and learn how to harness this understanding for anthropogenic processes, the Shafaat group will model the nickel-iron hydrogenases using a robust, protein-based scaffold. The proposed studies on the engineered enzymes are aimed at the elucidation of the reaction mechanisms along with the identification of the key factors along the entire protein contributing to catalysis. The research provides insight into how natural enzymes function across a range of length- and timescales. Graduate, undergraduate, and high-school students, including those from underserved communities, will be trained in state-of-the-art research techniques spanning biological, inorganic, physical, and analytical chemistry. This award also supports the development of lectures and hands-on exercises to train members of the broader bioinorganic community who attend the internationally recognized, biennial Penn State Bioinorganic Workshop. This project will be integrated with outreach programs spanning multiple age groups in greater Central Ohio. This project seeks to address critical knowledge gaps about the naturally occurring nickel-iron hydrogenases and related nickel-thiolate enzymes through the development and characterization of a model hydrogenase enzyme, namely nickel-substituted rubredoxin (NiRd). Prior work supported by the NSF in the Shafaat group has established that NiRd is a functional mimic of hydrogenase that exhibits high rates for hydrogen evolution. In the proposed work, spectroscopic techniques, including electron paramagnetic resonance, nuclear magnetic resonance, resonance Raman, and X-ray absorption, coupled with electrochemical methods, will be employed to obtain high-resolution mechanistic information and to reveal molecular-level contributors to hydrogen evolution activity. These studies will provide detailed insight into the layers of control exerted by the protein scaffold in both model and native metalloenzymes. The goals of the work are to identify correlations between enzymatic structure and function and to advance our understanding of biological hydrogen conversion. The research seeks also to identify design principles for highly active artificial metalloenzymes, with long-term implications for the development of sustainable catalysts for small molecule activation 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.