Two Methodologies in Pursuit of the Elucidation of Copper (II)—Centered Bioinorganic Chemistry

Author

William John Wagner, University of South Florida

Graduation Year

2009

Document Type

Thesis

Degree

M.S.

Degree Granting Department

Chemistry

Major Professor

Li-June Ming, Ph.D.

Co-Major Professor

Brian T. Livingston, Ph.D.

Committee Member

Steven H. Grossman, Ph.D.

Keywords

Aminopeptidase, Streptomyces, Copper, Moonlighting, Histatin

Abstract

Copper is a widely distributed transition metal in the earth's crust and has been adopted in a variety of biological systems. In many ways the biochemical usefulness of copper stems from its positive redox potential. This positive redox potential allows copper to assist in the movement of electrons. Copper ions can be found in natural systems as either Cu^I, Cu^II or Cu^III in part due to this redox potential. While Cu^II -centered biochemistry has been studied for years, mechanistic details in certain Cu^II -centered redox reactions remain unresolved. This study presents two methodologies for studying natural systems with known Cu^II -centered redox capabilities in order to better elucidate the mechanistic intricacies of Copper ion chemistry. The first method explored involves the promiscuous enzyme Streptomyces griseus aminopeptidase (SgAP) which although known primarily as a peptidase has been shown to oxidize catechol under near physiological conditions in vitro when its native Zn^II ions are replaced by Cu^II ions. Protein engineering techniques were utilized toward expression a functional recombinant enzyme in wild type and mutant forms. The goal was to utilize Site directed mutagenesis of residues in the active site to determine which residues are involved in both the hydrolysis and the oxidative activities of SgAP. The second methodology explored was the use of the N-terminus of Histatin-5, a naturally occurring peptide that is known to form complexes with Cu^II, as a model system to study Cu^II -centered oxidation chemistry. Metal-Peptide complexes are much more simplified model systems which use the same building blocks as proteins, but reduce the structure to the minimal functional unit necessary for activity. This in turn, simplifies the study of their catalytic chemistry as influences outside of the active region are greatly reduced. Furthermore, chemical synthesis of short peptides is easily performed and inexpensive in comparison to protein engineering, thus enabling further exploration, if deemed necessary, to be a feasible and economically viable possibility.

Scholar Commons Citation

Wagner, William John, "Two Methodologies in Pursuit of the Elucidation of Copper (II)—Centered Bioinorganic Chemistry" (2009). USF Tampa Graduate Theses and Dissertations.
https://digitalcommons.usf.edu/etd/73