Degree Type


Date of Award


Degree Name

Doctor of Philosophy


Biochemistry, Biophysics and Molecular Biology



First Advisor

Reuben J Peters


Terpenes consist of the most chemically diverse class of natural products and have an incredible scope of function and application. As such, understanding not only the vast array of uses for these molecules, but also how they are naturally produced is of great interest in biology and synthetic biology. Terpenes are produced in life from ubiquitous isoprene subunits which are concatenated at various lengths prior to being cyclized and functionalized to produce bioactive terpenoids. The biosynthesis of terpenoids therefore proceed down combinatorial networks of enzymatic pathways which account for the incredible diversity of products observed that originate from the common isoprene precursor. Diterpenes are a subclass of terpenes molecules which specifically consist of four isoprene subunits, or 20 carbons. The class of enzymes which catalyze the committed step in labdane related diterpenoid biosynthesis, class II diterpene cyclases (DTCs), from the universal twenty carbon precursor, geranylgeranyl diphosphate (GGPP), are therefore of great interest as they are the gatekeepers that determine which biosynthetic pathway this universal substrate will be guided down. DTCs catalyze their reaction via an acid-base catalysis mechanism wherein the substrate is first protonated forming a carbocation which can then be deprotonated in a variety of ways to yield a diverse set of cyclized products. By developing an understanding of the structure-function relationship of these enzymes, a concurrent understanding can be established on how the function of these enzymes can be predicted and engineered to result in higher capabilities in the ability to engineer organisms for desired phenotypes as well as the ability to produce these molecules biosynthetically. In this work, computational approaches, mutagenesis approaches, and analytical approached were used to both understand how evolution has attributed to the diverse product outcome of these enzymes and to rationally probe the active site residues of functionally characterized enzymes to alter product outcome. Our data has provided insights into the effects of key active site residues in determining product outcome, progress in method development of probing enzymes from an evolutionary perspective, and progress in the ability to rationally engineer DTCs for desired product outcome.


Copyright Owner

Cody Lemke



File Format


File Size

156 pages

Available for download on Friday, January 07, 2022