Date of Award
8-2025
Degree Name
Doctor of Philosophy
Department
Chemistry
First Advisor
Frederick Stull, Ph.D.
Second Advisor
Todd J. Barkman, Ph.D.
Third Advisor
David L. Huffman, Ph.D.
Fourth Advisor
Benjamin Koestler, Ph.D.
Keywords
Ancestral sequence resurrection, flavin amine oxidases, mechanistic enzymology, molecular evolution
Abstract
Flavoprotein amine oxidases (FAOs) are key enzymes in various kinds of metabolic pathways, mediating redox reactions via flavin cofactors. While most FAOs function as oxidases that readily reduce oxygen to hydrogen peroxide, a few outliers of the FAO family act as dehydrogenases that suppress the reaction with oxygen. The molecular basis for this divergence in function remains poorly understood.
The work presented in this study investigates the fundamental mechanisms of how flavincontaining enzymes activate or suppress their reaction with different electron acceptors to achieve high turnover rates. The overall body of work is divided into three different sections that (1) investigate the molecular basis and evolution of oxygen activation within a clade of enzymes where the majority of them are dehydrogenases, (2) compare and contrast the mechanism of two evolutionarily distant but functionally similar enzymes, (3) understand the evolutionary relationship between a group of dehydrogenases and their associated electron accepting partner proteins.
Through this work, the author successfully demonstrated that distal residues could modulate local active site dynamics and reactivity, providing the mechanistic basis for the evolutionary reemergence of oxidase function from within a larger clade of dehydrogenases. The author also highlighted how an independent evolutionary solution between two enzymes accomplished the same catalytic function. This work also presents evidence of a coevolutionary relationship between some flavin dehydrogenases and their associated cytochrome c partner proteins, which underscores the potential role of redox partner interactions in shaping flavoenzyme evolution.
Consequently, this work brings new insights into mechanistic enzymology, and how evolution of a certain activity can be driven by both local and distal amino acid substitutions that modulate protein dynamics of the active site architecture. These findings could be applied to other enzyme families and may provide new strategies for enzyme engineering for practical uses.
Access Setting
Dissertation-Open Access
Recommended Citation
Zhang, Zhiyao, "Mechanistic and Evolutionary Insights into a Group of Nicotine and Hydroxynicotine Degrading Flavoenzymes" (2025). Dissertations. 4195.
https://scholarworks.wmich.edu/dissertations/4195