Date of Graduation

5-2026

Document Type

Thesis

Degree Name

Bachelor of Science in Biology

Degree Level

Undergraduate

Department

Biological Sciences

Advisor/Mentor

Dr. Mack Ivey

Committee Member

Dr. Yuchun Du

Second Committee Member

Dr. Kate Walker

Third Committee Member

Dr. Mary Beth Long

Abstract

Clostridioides difficile is an opportunistic pathogen responsible for severe gastrointestinal infection. C. difficile-related infection is extremely prevalent in healthcare facilities, causing roughly 30,000 deaths annually in the United States alone. Its main mechanism of virulence is sporulation, which is directed by the SpoIIE protein. This makes SpoIIE an attractive target for the remedy and prevention of infections. The objective of this study was to analyze its structure using both lab techniques and artificial intelligence modeling. A truncated version of this protein, SpoII∆TM, was expressed and analyzed using optimal conditions previously determined in the lab. SDS-PAGE results and analyses confirmed the presence of the target protein in the sample. In order to obtain an adequate amount and purity of protein, future projects would need to conduct the experiments on a larger scale. They would then purify the amplified sample with Strep-tag protein purification and gain results from a cryogenic-electron microscopy collaborator. Multiple amino acid sequences were analyzed using AlphaFold Server to predict the 3-dimensional protein structure. These images revealed valuable information about the domains of SpoIIE and provided guidance for future studies. Using Foldseek, a protein database, the linker region of SpoIIE was compared to a HAMP region on a methyl-accepting chemotaxis protein. High homology between these domains provided evidence for the linker region’s role in dimerization and signal transduction.

Keywords

C. diff; SpoIIE

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