Hydrogen exchange equilibria in thiols.

Area of Science:

• Biochemistry
• Chemical Physics

Background:

• Peptides and small molecules undergo hydrogen-deuterium (H-D) exchange.
• Understanding H-D exchange mechanisms is crucial for studying molecular structure and dynamics.
• Radical-mediated reactions offer a pathway to probe H-D exchange.

Purpose of the Study:

• To investigate H-D exchange in various peptides including cysteine, cysteinyl-glycine, glutathione (GSH), phenylalanyl-cysteinyl-glycine, and histidyl-cysteinyl-glycine.
• To identify specific sites of H-D exchange under acidic and neutral conditions.
• To explore the influence of pH and molecular structure on radical-mediated H-D exchange.

Main Methods:

• Dissolving peptides in D(2)O (deuterated water) under acidic and neutral conditions.
• Utilizing a radical generator (2,2'-azobis(2-methylpropionamidine) dihydrochloride) and radical mediators (benzyl alcohol, 2-propanol).
• Employing Nuclear Magnetic Resonance (NMR) spectroscopy, including 2D NMR, to monitor H-D exchange and identify exchange sites.

Main Results:

• Significant H-D exchanges were observed in acidic media at specific positions within GSH, Phe-Cys-Gly, and His-Cys-Gly.
• In neutral media, exchanges were noted in Cys-Gly and GSH at distinct sites.
• 2D NMR revealed an unexpected rearrangement in GSH in acidic solution, consistent with thiolactone formation.

Conclusions:

• The sites of radical-mediated H-D exchange are highly dependent on peptide structure and pH.
• Electron density, particularly lone pairs, appears to facilitate the extent of exchange.
• Acidic conditions can induce structural rearrangements, such as thiolactone formation, in peptides containing cysteine and glycine residues.