Researcher ORCID Identifier

0000-0001-8655-4896

Graduation Year

2025

Document Type

Campus Only Senior Thesis

Degree Name

Bachelor of Arts

Department

Chemistry

Reader 1

Jane M. Liu

Reader 2

Matthew H. Sazinsky

Terms of Use & License Information

Terms of Use for work posted in Scholarship@Claremont.

Rights Information

© 2025 Giuliano Richetta

Abstract

To adapt to chemically complex nutritional environments, bacteria have evolved intricate mechanisms of genetic regulation. Vibrio cholerae, the bacteria responsible for cholera disease, can readily adapt its metabolism depending on which sugars are available in its immediate environment. In particular, V. cholerae has a phosphoenolpyruvate-dependent phosphotransferase system (PTS), a sugar-specific transport mechanism widely found across Gram-negative species. Across species, it has been observed that the PTS is a highly regulated system. Mannitol, a six-carbon alcohol sugar found at high concentrations in V. cholerae-abundant aquatic environments, is transported into the cell via mannitol-specific PTS transport. In V. cholerae, mannitol transport is mediated by the mtl operon, which encodes mannitol transporter MtlA and transcriptional regulator MtlR. Our understanding of gene- and protein-level pathways regulating the expression of the mtl operon remains limited: for instance, MtlR has been shown to downregulate transcription of mtlA by an unknown mechanism. This work explored the use of DNA Affinity Chromatography (DNA-AC) as a useful chemical biology tool to identify protein-DNA interactions in the mtl operon, specifically in its promoter region. The goal of this project was to optimize this pulldown method to understand V. cholerae gene regulation in response to its metabolic environment and elucidate the full suite of protein regulators acting at the mtl operon. My results show that DNA-AC allows for specific binding of proteins using purified protein lysate from V. cholerae; more work is needed to validate these findings before performing bottom-up proteomics by mass spectrometry to identify protein hits.

Download

This thesis is restricted to the Claremont Colleges current faculty, students, and staff.

Share

COinS