Structural determination of Zn²⁺, Cu²⁺, and Fe²⁺ complexed with glutathione by IRMPD spectroscopy and complimentary ab initio calculations

Glutathione is a biologically abundant and redox active tripeptide that serves to protect cells from oxidative stress and rid the body of toxic heavy metals. The present study examines the coordination complexes of glutathione (GSH) with metals that are central to redox processes in biology, Zn, Cu, and Fe, using infrared multiple photon dissociation (IRMPD) action spectroscopy with a free electron laser. For all three metals, a complex between the metal dication and deprotonated GSH was formed, M(GSH-H)⁺. The experimental IRMPD spectra were compared to scaled harmonic vibrational spectra calculated at the MP2/6-311+G(d,p) level of theory after thorough exploration of conformational space using a simulated annealing protocol. Interestingly, spectra calculated at the B3LYP or ωB97XD level do not match experiment as well. These findings offer the first gas-phase spectroscopic evidence for how the biologically relevant metal ions coordinate with glutathione. There are spectral features that are common to all three metals, however, noting the differences in the strengths of the common features between the three metals enables an assessment of the preference or specificity that each individual metal has for a given coordination site. Additionally, all three metals form structures where the deprotonated thiol of the cysteine side chain coordinates with the metal center, which is consistent with the involvement of the thiol site in biologically relevant redox chemistry.