Enzymatic cross-linking of proteins with tyrosinase

Tyrosinase (EC 1.14.18.1), usually known as the enzyme responsible for the enzymatic browning of fruits and vegetables, has been demonstrated to induce cross-linking of the whey proteins α-lactalbumin and β-lactoglobulin. The maximum degree of polymerisation for 1.5 mg/ml protein has been achieved at a tyrosinase activity of about 330 U/ml in the presence of a 2 mM concentration of caffeic acid. The pH optimum for the cross-linking reaction was detected at pH 4–5 for α-lactalbumin and β-lactoglobulin, in contrast to pH 7 for lysozyme, a non-whey protein, indicating that for the whey proteins the initial enzymatic oxidation of the phenolic compound is not the rate limiting step for the reaction. In contrast to β-lactoglobulin and lysozyme, for α-lactalbumin a direct cross-linking even without addition of caffeic acid has been proved. In this manner α-lactalbumin polymers with a molecular weight larger than 300 kDa have been produced by increasing the reaction temperature up to 50 °C. The results of the present study represent a basis for the creation of new high molecular weight proteins. Due to a different reaction mechanism and different points of linkage, tyrosinase-induced protein cross-linking may be a promising alternative to the already established use of transglutaminase for food application.