Altering the Regiospecificity of C6 Indole Prenyltransferase Enzymes Towards Drug Development

Author

Ahmed R. Aoun, Chapman UniversityFollow

Date of Award

Summer 8-2019

Document Type

Thesis

Degree Name

Master of Science (MS)

Department

Pharmaceutical Sciences

First Advisor

Sherif Elshahawi

Second Advisor

Keykavous Parang

Third Advisor

Aftab Ahmed

Abstract

Prenyltransferase (PT) biocatalysts are late-stage tailoring enzymes that modify natural products. PTs catalyze the attachment of prenyl moieties to natural product acceptors using pyrophosphate donors. This prenyl modification in small molecules leads to changes in structural and biological activities. Understanding the structural insights as well as the mechanisms by which PTs function allows us to utilize PTs as a unique approach towards drug development. PriB PT is an example of aromatic PTs that has been characterized previously as a L-tryptophan (L-Trp) C6 C-prenyltransferase (C6-C-PT). Analysis of PriB binding pocket, highlighted key residues that might play an important role in determining the regiospecificity of the enzyme and their mechanisms. Hence, we hypothesized that site-directed mutagenesis of one of these residues will alter the enzyme regiospecificity and/or its permissiveness leading to variation in the pharmacokinetics and biological activities of small molecules. Site-directed mutagenesis approach was used to engineer PriB PT as a model for C6-PTs and the purified mutant proteins were produced and purified. In vitro reactions of the purified engineered proteins have shown altered regiospecificity toward substrate acceptors when compared to wild type. The long-term goal for this study is to utilize the permissiveness of PriB enzyme for drug diversification and determine the activity of the mutant PriB enzymes. This study will also shed some light on the mechanistic insights of aromatic C6-PTs.

Creative Commons License

This work is licensed under a Creative Commons Attribution-Noncommercial-No Derivative Works 4.0 License.

Recommended Citation

Aoun AR. Altering the Regiospecificity of C6 Indole Prenyltransferase Enzymes Towards Drug Development. [master's thesis]. Irvine, CA: Chapman University; 2019. https://doi.org/10.36837/chapman.000087