Date of Award

Summer 2019

Document Type


Degree Name

Doctor of Philosophy (PhD)



First Advisor

Holz, Richard C.

Second Advisor

Gardinier, James R.

Third Advisor

St. Maurice, Martin


In industry, chemicals are typically synthesized through catalytically accelerated chemical processes. The use of biocatalysts is an increasingly important trend in achieving environmentally friendly chemical production processes. Nitrile Hydratases (NHases) are well-known biocatalysts that are used by several manufacturers such as Mitsubishi Rayon Corporation and Lonza for the conversion of acrylonitrile to acrylamide and 3-cyanopyridine to nicotinamide, respectively. Amides, in general, are used in polymer preparation, adhesive manufacturing, and the paper industry. Refrigeration is needed in the enzymatic hydration process on the industrial scale to maintain a low reaction temperature to stabilize the NHase enzyme. The use of whole cells instead of purified enzyme is one way to overcome this stability issue. However, the permeability of nitriles across the bacterial plasma membrane and cell wall is an important issue in order to get nitrile substrates into the cell and amide products excreted. The presence of unwanted enzymes in whole cells that can interfere with NHase activity, raw material, or product inhibition are also important considerations. Despite these issues, NHase shows great promise as a biocatalyst and, as a more detailed understanding of their catalytic mechanism and biological assembly processes emerge, improved biocatalyst designs that are more efficient, stable, and selective will advance their industrial use. The goal of the dissertation is to: study the formation of Fe-Type α(ɛ) protein complex, investigate the cellular maturation of Fe-Type nitrile hydratase, and discover intermediates along the catalytic cycle of NHases in-order to gain additional insight into the bio-assembly and catalytic mechanism of NHase enzymes.

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