Haïssat S, Marchal E, Montagne P, Humbert G, Béné M C, Faure G, Linden G
Laboratory of Applied Biochemistry, Faculty of Sciences, Vandoeuvre les Nancy, France.
Anal Biochem. 1994 Nov 1;222(2):472-8. doi: 10.1006/abio.1994.1519.
The binding of bovine plasminogen to whole casein, alpha s-casein, beta-casein, and kappa-casein is responsible for the progressive proteolysis of milk and dairy products. A sensitive and accurate microparticle-enhanced nephelometric immunoassay was developed to measure free plasminogen after interaction between bovine plasminogen and caseins and the quantitative parameters of plasminogen/casein binding were established. Two classes of binding sites for plasminogen were found in this study on each of the investigated caseins. Their dissociation constants (Kd) 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 (Kd = 32 nM) and the other of lower affinity (Kd > 370 nM). The number of both binding sites per casein monomer was low (0.04 to 0.53). The great propensity of casein monomers to self-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.
牛纤溶酶原与全酪蛋白、αs-酪蛋白、β-酪蛋白和κ-酪蛋白的结合是导致牛奶和乳制品逐步蛋白水解的原因。开发了一种灵敏且准确的微粒增强散射比浊免疫测定法，用于测定牛纤溶酶原与酪蛋白相互作用后的游离纤溶酶原，并确定了纤溶酶原/酪蛋白结合的定量参数。本研究在每种被研究的酪蛋白上发现了两类纤溶酶原结合位点。通过在pH 6.6条件下改变纤溶酶原浓度并进行Scatchard分析，确定了它们的解离常数（Kd）。这两个结合位点似乎一个具有高亲和力（Kd = 32 nM），另一个具有较低亲和力（Kd > 370 nM）。每个酪蛋白单体上这两个结合位点的数量都很少（0.04至0.53）。酪蛋白单体极易在同聚物中自缔合（纤溶酶原结合位点可能被隐藏），并以全酪蛋白胶粒的形式存在于牛乳中的共聚物中，这就导致计算得出每个单体的结合位点<1。
