Gonias S L, Pizzo S V
J Biol Chem. 1981 Dec 10;256(23):12478-84.
The plasma protease inhibitor alpha 2-macroglobulin (alpha 2M) was reacted in vitro with cis-dichlorodiamineplastinum(II) (cis-DDP). Following the reaction, alpha 2M demonstrated a significantly decreased ability to bind trypsin as determined by esterase activity assays in the presence of soybean trypsin inhibitor and studies with radiolabeled trypsin. Inactivation of alpha 2M by cis-DDP was not associated with a conversion to the "fast" electrophoretic form, as determined on nondenaturing gels, in contrast to the inactivation of alpha 2M by proteases and certain amine salts. The extent of reaction increased with the elevation of temperature within the thermal stability range of the protein; however, variation of pH within the range 6.82-8.55 had little effect. Binding of [14C]methylamine to alpha 2M was not affected by cis-DDP. The conformational change, however, which normally accompanies this reaction did not occur. It is concluded that the alpha 2M thiolesters are most likely not reactive sites for cis-DDP. cis-DDP-treated alpha 2M failed to dissociate into quarter subunits under denaturing and reducing conditions, suggesting cross-linking of subunits. This cross-linking may be responsible for locking the alpha 2M quarternary structure into the "slow conformation."
血浆蛋白酶抑制剂α2-巨球蛋白（α2M）在体外与顺二氯二氨铂（II）（顺铂，cis-DDP）发生反应。反应后，通过在大豆胰蛋白酶抑制剂存在下的酯酶活性测定以及用放射性标记胰蛋白酶进行的研究发现，α2M结合胰蛋白酶的能力显著下降。与蛋白酶和某些胺盐使α2M失活不同，非变性凝胶电泳结果显示，顺铂使α2M失活与转化为“快”电泳形式无关。在蛋白质热稳定性范围内，反应程度随温度升高而增加；然而，在6.82 - 8.55范围内改变pH值影响不大。[14C]甲胺与α2M的结合不受顺铂影响。然而，通常伴随此反应的构象变化并未发生。得出的结论是，α2M硫酯很可能不是顺铂的反应位点。在变性和还原条件下，经顺铂处理的α2M无法解离成四分之一亚基，提示亚基发生了交联。这种交联可能是导致α2M四级结构锁定为“慢构象”的原因。
