Date of Award
Doctor of Philosophy (PhD)
Biochemistry & Molecular Biophysics
Peter L. Dutton
In nature, oxidoreductase proteins are responsible for many enzymatic processes critical to life. These proteins often rely on the presence ofÂ non-proteinaceous cofactorsÂ to take part in the enzymatic function. Â The most common,Â central to my thesis,Â is heme B. Â Depending on the protein environment, this cofactor can take part in functions as diverse as electron transfer (cytochromes), oxygen transport (hemoglobins), oxygen reduction (oxidases), carbon-hydroxylation (oxygenases), and superoxide production (NADH oxidase).
In natural oxidoreductases, determination of the course and rates of heme-protein association, what barriers are encountered, what affinity is achieved, and what are the oxidation-reduction potentials,Â is criticalÂ forÂ understanding the rules ofÂ assembly and function of the different activities performed. In the growing field of research attempting to make man-made oxidoreductases, the same understanding is required for progress to be made towardÂ construction of novel enzymes. However, this understanding is still out of reach in natural oxidoreductasesÂ because of the immenseÂ complexity of natural proteins, while for man-made designs progress has only recently reached a point where an in-depth systematic study can be contemplated.Â
My thesis states: Simple non-natural proteins (maquettes) designed from first principles to ligate heme, can be used toÂ uncover the factors derived from theÂ oligomeric andÂ structural state of related maquette and also derived from porphyrin variants of heme B,Â that govern rates of Â incorporation and ligation ofÂ heme BÂ into a maquette.Â Maquettes are ideal platforms to demonstrate what aspects of a protein govern heme redox potentials, a key parameter underlying the diversity of hemoprotein functions. Â
The findings from my work provide the first views of heme and maquette assembly: spontaneous, rapid and with high affinity association. They also provide a foundation for understanding what controls redox potentials of the heme and perspective on this control. The work offers insight into similar processes in natural oxidoreductases,Â but the concepts and principles uncovered in this thesis will be vital in the development ofÂ novel functions applied in man-made applications in vitro and in vivo.
Solomon, Lee Andrew, "The Physical Chemistry Underlying the Assembly and Midpoint Potential Control in a Series of Designed Protein-Maquettes" (2013). Publicly Accessible Penn Dissertations. 802.