Redox reactions of and transformation between cysteine-mercury thiolate and cystine in metallothioneins adsorbed at a thin mercury film electrode

Voltammetric studies of rabbit liver metallothioneins (MTs) adsorbed onto t hin mercury films preformed onto glassy carbon electrodes were carried out in MT-free phosphate buffers in an attempt to shed light on the possible re dox-modulated MT metal transfer processes. The redox behavior of the surfac e-confined MTs was studied by cyclic voltammetry and differential pulse vol tammetry, and the amount of MT adsorption was quantified by a flow-injectio n quartz crystal microbalance. Two reversible redox waves, with E, values a t -0.63 and -0.91 V, respectively, were observed for the first time. These values and the overall voltammetric characteristics were found to be remark ably analogous to those of cysteine adsorbates at thin mercury films. The p eak at E-p = -0.63 V is attributable to the reduction of the Cys-mercury th iolates formed between the adsorbed MTs and the mercury electrode, whereas that at E-pc = -0.91 V is assigned to the reduction of Cys-Cys (cystine ana logue) present in the portion of the MT adsorbate that is not in direct con tact with the mercury film. The two redox waves were observed to be interch angeable through preelectrolysis at a negative potential (e.g., -1.1 V) to reduce the MT adsorbate, or at a more positive potential (e.g., -0.1 V) to oxidize the adsorbed MTs. On the basis of these voltammetric results, we pr oposed a simple schematic to elucidate the redox reactions of and the trans formation between the cysteine-mercury thiolates and cystines that are pres ent in the :MT adsorbates under different electrochemical redox conditions. Since the electrochemical reduction of the cystine analogues in the MT ads orbates to the corresponding cysteines (a process facilitating metal comple xation) and the reoxidation of the cysteine residues back to cystine analog ues (a process causing metal release) are both reversible, it appears that the metal transfer between MT and a substrate might accompany the variation of the redox states of the MT-metal complexes. Our voltammetric studies of MTs thus provide supportive evidence for the mechanism of the MT metal tra nsfer in cytoplasmic milieu proposed by Vallee and co-workers (Maret, W.; V allee, B. L. Proc. Nail. Acad. Sci. U.S.A. 1998, 95, 3478-3482).