Roossien F F, van Es-Spiekman W, Robillard G T
FEBS Lett. 1986 Feb 17;196(2):284-90. doi: 10.1016/0014-5793(86)80264-2.
The occurrence of intermolecular dithiols on EIImtl has been studied with a number of thiol-specific cross-linking reagents. The reaction of EIImtl with bifunctional maleimide derivatives inactivates the enzyme. At the same time the enzyme is irreversibly cross-linked to a dimeric species. Under optimal conditions 50% of the protein is cross-linked upon reaction with the dimaleimides. The enzyme is also cross-linked under oxidizing conditions in the presence of CuCl2, presumably by oxidizing an intermolecular dithiol to a disulfide. This oxidation can be reversed by the addition of the reducing agent dithiothreitol. The reaction of phosphorylated EIImtl with the same sulfhydryl-specific bifunctional reagents does not lead to any cross-linked product. The results are discussed in terms of the association state of the purified protein and the distribution of its thiol groups.
已使用多种硫醇特异性交联试剂研究了EIImtl上分子间二硫醇的出现情况。EIImtl与双功能马来酰亚胺衍生物的反应使该酶失活。同时，该酶不可逆地交联成二聚体形式。在最佳条件下，与双马来酰亚胺反应时50%的蛋白质会发生交联。在CuCl2存在的氧化条件下，该酶也会发生交联，推测是通过将分子间二硫醇氧化为二硫键实现的。加入还原剂二硫苏糖醇可使这种氧化反应逆转。磷酸化的EIImtl与相同的巯基特异性双功能试剂反应不会产生任何交联产物。根据纯化蛋白质的缔合状态及其硫醇基团的分布对结果进行了讨论。
