Quantification of Oxidized Metallothionein by a Cd-Saturation Method

One of the most prominent characteristics of metallothionein (MT) is its high cysteine content of 30%. The sulfhydryl groups, normally involved in metal binding, render the protein particularly sensitive towards oxidation. Oxidation of sulfhydryl groups in MT may occur in vivo and in vitro, either intra- or intermolecularly, the latter leading to dimeric or polymeric forms of MT. Thiol oxidation and oligomerization of MT may be induced by various radicals (1-3 ) and free metal ions (4 ). The sensitivity to oxidation of thiol groups in MT to a great extent depends on the metals that are bound to the protein. Particularly the Cu-containing MT easily polymerizes (3,5 ). Accordingly, Cu-rich polymerized MT has been isolated from the livers of newborn animals (6 ) and human fetuses (7,8 ). Insoluble polymerized forms of MT were shown to be mainly located in the heavy lysosomal fraction of the liver (6 ). In agreement, elevated levels of Cu associated with oxidized forms of MT have been described in lysosomes of patients with Wilson disease and dogs with inherited Cu-toxicosis (9-12 ). More recently, polymeric MT has been also detected in lysosomes of hepatocytes and Kupffer cells from LEC rats, the rodent model for Wilson disease (13 ).