Date Thesis Awarded

5-2015

Access Type

Honors Thesis -- Access Restricted On-Campus Only

Degree Name

Bachelors of Science (BS)

Department

Chemistry

Advisor

Lisa M. Landino

Committee Members

Christopher J. Abelt

Gary W. Rice

Pamela S. Hunt

Abstract

Protein oxidation and repair is a physiological process that has been implicated in the pathology of diseases such as Alzheimer’s and in the normal aging process. S-glutathiolation, the process by which glutathione reacts at equilibrium with a protein and prevents the protein from undergoing further oxidation, is a critical repair mechanism for oxidized proteins.

In this thesis, we use the technique of fluorescence spectroscopy to develop a methodology to visualize S-glutathiolation in vitro. We report the spectroscopic properties of previously synthesized glutathione derivatives that have successfully labeled proteins during periods of oxidation. We also report the results of labeling BSA and papain, two proteins that contain one cysteine, at their cysteine and amine residues as well as the effect of unfolding on the dansyl emission wavelength for the cysteine-labeled proteins. In addition, we report the emission wavelengths from four proteins: BSA, papain, CK, and GAPDH, labeled with D-GSH in the presence of H₂O₂.

The results from the experiments herein demonstrate the usefulness of this methodology. Oxidative labeling of proteins with D-GSH results in an emission wavelength that varies between proteins and is lower than the D-GSH emission wavelength. While this proves to be a promising result, the model is only tested with proteins containing up to four cysteines and the labeling mechanism for multi-cysteine proteins is not currently understood. Further refinement is required before implementation.

Recommended Citation

Daniels, Jacob, "Novel methodologies to study protein S-glutathiolation using fluorescence spectroscopy" (2015). Undergraduate Honors Theses. William & Mary. Paper 157.
https://scholarworks.wm.edu/honorstheses/157