Inhibition of plasmin by beta-lactoglobulin using casein and a synthetic substrate.

Bovine plasmin (EC 3.4.21.7) activity was measured on H-D-valyl-L-leucyl-L-lysyl-4-nitroanilide and acid casein in the presence of native and heat-denatured beta-lactoglobulin (denatured at 100 degrees C for 15 min before being mixed with plasmin solutions). Native or denatured beta-lactoglobulin was then heated with plasmin at 60 degrees C for 15 min. Enzyme activity again was estimated after this mild heat treatment. Native and denatured beta-lactoglobulin inhibited the action of plasmin on H-D-valyl-L-leucyl-L-lysyl-4-nitroanilide and casein. The mild heat treatment (60 degrees C for 15 min) caused stronger inhibition of the activity of plasmin against casein and the synthetic substrate. For H-D-valyl-L-leucyl-L-lysyl-4-nitroanilide, inhibition was competitive in unheated mixtures, but heating beta-lactoglobulin with plasmin changed inhibition type to mixed. This change suggests a heat-dependent interaction between plasmin and beta-lactoglobulin. Native beta-lactoglobulin was more inhibitory of plasmin's action against casein than was denatured beta-lactoglobulin. The converse was observed when plasmin activity was measured with the synthetic substrate.