QUANTITATIVE CHARACTERIZATION OF BOVINE PLASMINOGEN BINDING TO CASEINS

The binding of bovine plasminogen to whole casein, alpha(s)-casein, be ta-casein, and kappa-casein is responsible for the progressive proteol ysis of milk and dairy products. A sensitive and accurate microparticl e-enhanced nephelometric immunoassay was developed to measure free pla sminogen after interaction between bovine plasminogen and caseins and the quantitative parameters of plasminogen/casein binding were establi shed. Two classes of binding sites for plasminogen were found in this study on each of the investigated caseins. Their dissociation constant s (K-d) were determined by varying the plasminogen concentration at pH 6.6 and performing Scatchard analysis. The two binding sites appeared to be one of high affinity (K-d = 32 nM) and the other of lower affin ity (K-d > 370 nM). The number of both binding sites per casein monome r was low (0.04 to 0.53). The great propensity of casein monomers to s elf-associate in homopolymers where plasminogen binding sites could be hidden and in copolymers present in bovine milk in the form of whole casein micelles accounts for calculated binding sites <1 per monomer. (C) 1994 Academic Press, Inc.