Characterization of the [FeFe]-hydrogenase : toward understanding and implementing biohydrogen production
Swanson, Kevin Daniel.
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Hydrogen may provide an avenue for a clean renewable fuel source, yet the methods to produce hydrogen are either extremely energy intensive, rely on fossil fuels, or require expensive noble metal catalysts. Biology may hold the keys necessary to unlocking new technologies that could change how hydrogen is produced. Microbial processes also produce hydrogen and harbor enzymes that carryout the reversible reduction of protons to hydrogen gas. These enzymes are capable of producing hydrogen at high rates comparable to platinum catalysts, but biological hydrogen catalysts can produce hydrogen using abundant elements carbon, oxygen, nitrogen, sulfur, iron, nickel and selenium. Biological hydrogen catalysts are termed hydrogenases, and though hydrogenases use abundant elements they are extraordinarily complex. This has made it difficult to construct model complexes using inorganic synthesis that can replicate the activities of their biological counterparts. One way to circumvent this problem is to use microbial hydrogen production and let microbes produce and maintain these enzymes inside a cell. Microbial hydrogen production also has the added benefit that hydrogen production could be engineered to connect with other metabolic processes such as photosynthesis and fermentation. Engineering microbes for hydrogen production could eventually allow for the production of hydrogen using inexpensive energy inputs such as solar energy or waste materials. Yet, there are many barriers that need to be overcome in order to engineer a robust microbial organism. One of the primary difficulties of developing this technology has been the oxygen sensitivity of hydrogenases. Hydrogenases when exposed to atmospheric concentrations of oxygen either completely inactivate or their rates are significantly slowed. To engineer a hydrogenase that is more amenable for microbial hydrogen production, the optimization of expressing and purifying hydrogenase enzymes has been developed. Methodologies have been developed to characterize how oxygen inactivates hydrogenase enzymes, and a new methodology has been explored to help find novel hydrogenase gene sequences that may help in engineering oxygen tolerant enzymes.