Calcium- and cysteine-participatory oxidative formation of albumin-copper complex

An albumin-copper-cysteine complex that has been shown to be formed in serum when cupric ion is injected intravenously into rats was reconstituted in vitro with mercaptalbumin, cupric ion, and cysteine in the presence of calcium ions. Mercaptalbumin was separated from fresh rat serum on a GS-520P column and the protein was incubated with an equimolar amount of cupric ions, which resulted in the formation of an albumin-copper complex. However, the albumin-copper complex did not form the albumin-copper-cysteine complex in the presence of cysteine at any of the molar ratios of cysteine to mercaptalbumin examined. Albumin-copper-cysteine complex was formed only when a more than equimolar amount of calcium to mercaptalbumin was present in the incubation mixture. The stoichiometric relationship revealed for the formation of albumin-copper-cysteine complex is mercaptalbumin: copper:cysteine:calcium = 1:1:2:1. One molar equivalent of cysteine was incorporated into the complex through disulfide bonds and another molar equivalent of cysteine was present unchanged after the reaction, suggesting that the latter cysteine participated in maintaining a specific conformation of albumin during formation of the complex. The overall reaction was oxidative and oxygen enhanced the reaction.