The Chemistry of Life Processes Program in the Chemistry Division is funding Dr. Andrew J. Wommack from High Point University to examine the functional and structural consequences of sulfur-sulfur or disulfide bonds in proteins. The disulfide bond serves important roles in biology ranging from structural effects in proteins to cell signaling and control of metabolic pathways. This sulfur-sulfur chemical bond can be found in opened or closed states under different biological conditions, such as during different physiological stresses or within unique cellular locations. To study the functional impact of the disulfide bond in a permanently closed state, one sulfur atom is replaced by a carbon atom to generate an alternative bond that is locked in place under all biologically relevant conditions. In addition to inventing new chemical methods for exploring how disulfide bond status affects biological processes, this project directly impacts the training of undergraduate research students and also facilitates development of new engaging materials for the undergraduate science curriculum.
Reactive oxygen/nitrogen species (ROS/RNS) are present in cells at various concentrations, depending on cell type, subcellular location, and physiological state. As an evolved response to the levels of ROS/RNS in oxidative metabolism, glutathione (GSH) is installed as a protected, but redox-reversible, covalent change to solvent-accessible cysteine thiols. By replacing the internal cysteine of GSH with an alkene-containing side chain, this modified GSH is irreversibly attached to solvent-accessible cysteines with optimized methods for thiol-ene coupling (TEC) in aqueous media. Initial targets include peroxiredoxins 1 & 2 (Prx1 & Prx2) and sulfiredoxin 1 (Srx1), where glutathionylation is critically involved in ROS/RNS equilibria and signaling. Mutagenesis of existing cysteine residues, followed by TEC reaction with the alkene-containing GSH, is assessed with LC-MS analysis, gel-filtration chromatography, spectrophotometry, analytical ultracentrifugation, and x-ray crystallography to explore how glutathionylation affects protein function and structure. With the developed TEC methodology, this project further contributes to the fundamental understanding of glutathionylation as a response to ROS/RNS.
This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.
