Characterizing Factors That Influence Intracellular Thiol-Disulfide Equilibrium

<p>Thiol-disulfide balance is critical for the proper functioning of many proteins. Reduced thiol residues can aid in cofactor binding and catalysis, while disulfide bonds are often required for protein folding and stability. Therefore, oxidation of critical cysteine residues can either activate or inactivate a protein based on its function in the cell. The on-going challenge for aerobic organisms is to maintain proteins in their functional redox state while promoting redox processes required for cell growth.</p> <p>The first part of this thesis focuses on determining the role that the oxidoreductase human glutaredoxin 1(hGrx1) plays in reduction of the critical disulfide bond in human superoxide dismutase 1 (hSOD1). hSOD1 is normally a protective enzyme that detoxifies superoxide (O2*). However, mutations in hSOD1 can cause amyotrophic lateral sclerosis (ALS or Lou Gehrig's disease), a common neurodegenerative disease. Our results indicate that hGrx1 uses a monothiol mechanism to preferentially reduce the intramolecular disulfide bond in ALS mutant forms of hSOD1. In addition, both in vivo and in vitro studies suggest that the intracellular redox state of hSOD1 may be controlled by kinetics rather than thermodynamics.</p> <p>The second part of this thesis focuses on understanding the factors that influence the mitochondrial GSH:GSSG balance by using genetically engineered in vivo fluorescence redox sensors, rxYFP and roGFP2 (yellow and green fluorescence proteins). This project focuses on how specific oxidoreductases and their assembly factors influence the subcellular GSH:GSSG redox potential. In one study, deletion of the mitochondrial iron homeostasis factor Mtm1 was found to cause a large oxidative shift in the GSH:GSSG redox potential in both the mitochondrial intermembrane space (IMS) and matrix. In contrast, deletion of mitochondrial matrix superoxide dismutase 2 has a smaller effect on the mitochondrial GSH:GSSG redox potential under similar conditions. In another study, the IMS-localized cytochrome c assembly factor Cyc2 was found to cause a reductive shift in the GSH:GSSG IMS redox state. Overall, these studies are aimed at defining factors that influence the subcellular GSH:GSSG redox state, which in turn affects thiol-disulfide equilibrium.</p>

Name thesis:degree_name

Ph.D.

Level thesis:degree_level

Campus Access Dissertation

Discipline thesis:degree_discipline

Chemistry and Biochemistry

Year

2010

Author dc:creator

• Bouldin, Samantha Diane

Contributors dc:contributor

• Caryn E. Outten

• Chemistry
• Physical Sciences and Mathematics
• Bouldin_sc_0202A_10735.pdf

Statement dc:rights

• © 2010, Samantha Diane Bouldin

Repository record dc:identifier

https://scholarcommons.sc.edu/etd/143

OAI identifier oai:identifier

oai:scholarcommons.sc.edu:etd-1144