Krebs G, Guinand S, Bréda C
C R Acad Hebd Seances Acad Sci D. 1978 Apr 24;286(16):1219-22.
A comparative study of the dissociation into subunits of Porcine alpha2 M, either native or bound to trypsin (Tn), has been carried out in order to determine the modifications of the alpha2 M structure due to the formation of the Tn-alpha2 M complex. Analytical ultra-centrifugation at pH 3.5 shows that the dissociation is smaller when alpha 2 M is bound to trypsin. Electrophoresis in 4% polyacrylamide gels, in presence of 0.1% SDS, of alpha2 M and Tn-alpha2 M incubated in 1% SDS leads to the same conclusion; the enzyme must stabilize the quaternay structure of alpha2 M. In presence of SDS + beta-mercaptoethanol, only a molecular weight (M.W.) 200,000 band is revealed in electrophoresis pattern of native alpha2 M. In the case of reduced Tn-alpha2 M, some other bands of M.W. 100,000, 50,000, 30,000 appear. When trypsin is inactivated by TLCK 100,000 M.W. band is present, accompanied by the 200,000 M.W. band whose intensity is function of the alpha2 M concentration. The 100,000 M.W. band appears therefore characteristic of the formation of the complex which must imply a proteolytic cleavage in the middle of the 100,000 polypeptidic chain of alpha2 M. A model of the complex is proposed in which the enzyme forms a proteic bridge between the two halves of the alpha2 M molecule.
为了确定由于形成胰蛋白酶-α2M复合物而导致的α2M结构的改变，对天然或与胰蛋白酶（Tn）结合的猪α2M亚基解离进行了比较研究。在pH 3.5下进行的分析超速离心表明，当α2M与胰蛋白酶结合时，解离程度较小。在1% SDS中孵育的α2M和Tn-α2M在含有0.1% SDS的4%聚丙烯酰胺凝胶中进行电泳，得出了相同的结论；该酶必须稳定α2M的四级结构。在SDS + β-巯基乙醇存在的情况下，天然α2M的电泳图谱中仅显示出一条分子量（M.W.）为200,000的条带。在还原的Tn-α2M的情况下，出现了一些其他分子量为100,000、50,000、30,000的条带。当胰蛋白酶被TLCK灭活时，存在100,000 M.W.的条带，同时伴有200,000 M.W.的条带，其强度是α2M浓度的函数。因此，100,000 M.W.的条带似乎是复合物形成的特征，这必然意味着在α2M的100,000多肽链中间发生了蛋白水解切割。提出了一个复合物模型，其中酶在α2M分子的两半之间形成了一个蛋白质桥。
