DIMERIC ALPHA(S2)-CASEIN, A NOVEL MATRIX FOR TISSUE-TYPE PLASMINOGEN-ACTIVATOR CATALYZED PLASMINOGEN ACTIVATION

In previous studies, we demonstrated a differential distribution of pl asminogen, tissue-type plasminogen activator (t-PA), and urokinase-typ e plasminogen activator (u-PA) between Various milk fractions. Plasmin ogen and t-PA were mainly confined to the casein micelle fraction, whe reas u-PA was restricted to the cell fraction. These studies also show ed that the differential distribution of these components was caused b y specific binding components in milk. The u-PA binding component in t he cell fraction was identified as the u-PA receptor. The t-PA binding components of bovine casein micelles were identified as dimeric alpha (s2)-casein and multimeric forms of K-casein. In order to further expl ore the action of the plasminogen activation system that influences en dogenous degradation of proteins in bovine milk, we evaluated the effe ct of individual bovine caseins on the rate of plasminogen activation by t-PA. Using a coupled peptidyl anilide plasminogen activator assay, we found that dimeric alpha(s2)-casein induced a concentration-depend ent enhancement of the t-PA-catalyzed plasminogen activation. On a wei ght basis, the stimulating effect of dimeric alpha(s2)-casein was comp arable to that of cyanogen bromide cleaved fibrinogen fragments. In a direct catalytic assay with a peptidyl anilide substrate, dimeric alph a(s2)-casein had no effect on t-PA activity. Ligand blotting experimen ts showed binding of both plasminogen and t-PA to the dimeric alpha(s2 )-casein. The combined results from the binding experiments and the en zymatic assays suggest that the observed enhancement in plasminogen ac tivation is mediated via formation of a ternary complex between t-PA, plasminogen, and dimeric alpha(s2)-casein. We further investigated the binding specificity of t-PA towards dimeric alpha(s2)-casein using t- PA deletion mutants and monoclonal antibodies. The results were compat ible with the binding being mediated by kringle 2 and the finger domai n of t-PA. These findings suggest that casein, analogous to fibrin clo ts in the intravascular space, serve as a molecular matrix for t-PA-ca talyzed plasminogen activation. Published by Elsevier Science B.V.