Biosynthesis and Reactivity of the Active Site of the FeFe Hydrogenases

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Project Summary Hydrogenase enzymes are pervasive, being found in bacteria, archaea, and some higher organisms. These enzymes are hosted by some pathogens, often in anaerobic environments including the human gut. The hydrogenases mediate the most fundamental chemical reaction: the interconversion of H2 with protons and reducing equivalents. The enzymes are structurally exceptional with an array of distinctive cofactors, especially the site of H2 binding and release. Interest in such enzymes stems from three angles: the possibility that some pathogens could be controlled rationally, the excitement about their unusual structures, and the commercial implications of hydrogen production/oxidation in the context of fuel cells. Two major classes of hydrogenases exist, [NiFe]- and [FeFe]-hydrogenases. This project is almost exclusively focused on the latter. More specifically, this project aims to elucidate the biosynthesis of the active site of [FeFe] enzymes, the faster hydrogenase and the one most amenable to development for other applications. This project is timely because we have just defined the sequence by which the three maturase enzymes build the active site. In parallel with their unusual structures, the construction (biosynthesis) of the active site proceeds unusually. The first subproject aims to make the first Fe-containing intermediate, "Compound B". The next two projects tackle how B is converted to an inorganic Fe-S-CN-CO monomer. The fourth project examines the coupling of this monomer to give an inorganic dimer. The final and fifth project examines the retrofitting of this Fe2 entity with an organic cofactor. In this program collateral projects address allied themes of still broader interest. One involves expanding our knowledge of iron complexes of amino acids. Another contributes to the biosynthesis of [NiFe]-hydrogenases. One spin-off project critically examines the premises of the Iron-Sulfur Theory of the origin of life by examination of the first Fe-S-CN-CO complexes.