Date of Award

6-2022

Document Type

Restricted (Opt-Out)

Degree Name

Bachelor of Science

Department

Biology

First Advisor

Kristin Fox

Second Advisor

J. Stephen Horton

Keywords

protein, metacaspase, expression, biochemistry, biology, western blot, mutagenesis

Abstract

Apoptosis is a form of programmed cell death (PCD), a critical process among the kingdoms of life especially in the processes of development, aging, and responses to stress. Fungi utilize metacaspases, cysteine-dependent proteolytic enzymes, as their primary apoptotic protein. Therefore, studying the proteolytic mechanisms of metacaspases can potentially lead to a better understanding of opportunistic fungal infections. There are three types of metacaspases categorized accordingly as Type I, Type II, and Type III. These types each have various structures and functions in apoptosis. Each metacaspase has a p20-like domain containing the catalytic active site for protease activity and a p10-like domain important for regulation of activity. The subject of this research is Schizophyllum commune (S. commune), a basidiomycete fungus. Previous literature has centered on Type I metacaspases, but this project focuses primarily on Type II metacaspases. There are 6 Type II S. commune metacaspases labeled a-f. All of these types contain a linker region between their p20 and p10 domains. In this thesis, I focus on expressing the Type IIa metacaspase from S. commune in order to develop an in-depth characterization and compare it to the previous research on S. commune Type I metacaspases. Expression of full-length ScMCA-IIa was unsuccessful while mutant ScMCA-IIa expression was successful. Therefore, we concluded nonconserved residues must be deleted from the metacaspase DNA sequence prior to proper expression. Expression is necessary before any further steps. Future work includes protein purification, characterization of the protein, and comparisons to Type I metacaspases.

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