Mechanism of decreased heat-induced activation of plasminogen in the presence of cysteine

The mechanism by which activation of plasminogen (PG) is reduced by heating in the presence of free sulphydryl groups was characterized, and the effect on PG structure was evaluated. Activity assays were performed on PG or milk containing 3–4 cysteine concentrations and heated at three temperatures. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis was carried out in one and two dimensions to visualize changes in PG. Activity assays with other sulphydryl- and non-sulphydryl-containing reducing agents, β-mercaptoethanol, glutathione and sodium borohydride, were performed. In heated PG samples, increasing cysteine concentration decreased activation of PG; temperature had less affect. Electrophoresis results confirmed the presence of disulphide-bonded PG polymers and the reversibility of polymerization. Activity assays with the other reducing agents confirmed that the mechanism was related to free sulphydryl groups. Activation of PG in milk can be partially controlled by addition of cysteine before pasteurization, due to a sulphydryl-reducing and polymerization mechanism.