Ethoxyformylation and photooxidation of histidines in transferrins.
Ethoxyformylation and photooxidation of histidines in transferrins.
复制标题
转铁蛋白中组氨酸的乙氧基甲酰化和光氧化。
作者：
T. Rogers;R. Gold;R. Feeney
利用两种不同的反应研究了卵转铁蛋白和人血清转铁蛋白中组氨酸的化学反应性。在染料敏化的光氧化的卵转铁蛋白和乙氧基化的人血清转铁蛋白和卵转铁蛋白，组氨酸和铁结合活性的损失进行了观察。两种脱辅基蛋白中的所有组氨酸都可以通过使用170至400摩尔过量的试剂而被乙氧基甲酰化，导致活性完全丧失。人血清转铁蛋白的组氨酸对试剂的反应性比卵转铁蛋白的反应性大。每个铁的结合保护两个组氨酸免受乙氧基化物的侵害，在这两种情况下，蛋白质都保持完全活性。组氨酸和铁结合活性的一阶损失时，观察到卵转铁蛋白照射在亚甲蓝的存在下。一级速率的比较表明，每个结合位点的两个组氨酸的损失占蛋白质的失活。然而，铁结合并没有保护卵转铁蛋白的光灭活预期。这两种修饰方法的结果表明：（1）组氨酸是铁结合活性所必需的。(2)每个结合位点有两个必需的组氨酸。本工作证明了在研究组氨酸在蛋白质中的功能作用时，使用乙氧基化和光氧化两种修饰反应的优点。
The chemical reactivity of histidines in ovotransferrin and human serum transferrin was studied utilizing two different reactions. Upon dye-sensitized photooxidation of ovotransferrin and ethoxyformylation of human serum transferrin and ovotransferrin, losses in histidine and iron-binding activity were observed. All of the histidines in both apoproteins could be ethoxyformylated by the use of 170 to 400 molar excesses of reagent resulting in complete loss in activity. The histidines of human serum transferrin showed a greater reactivity toward the reagent than did those of ovotransferrin. The binding of each iron protected two histidines from ethoxyformylation, and in both cases the proteins remained completely active. First-order losses in histidine and iron-binding activity were observed when ovotransferrin was irradiated in the presence of methylene blue. Comparison of the first-order rates indicates the loss of two histidines per binding site accounts for the inactivation of the protein. However, iron binding did not protect ovotransferrin from photoinactivation as expected. Evidence from both modification technqiues indicates: (1) Histidines are essential for iron-binding activity. (2) There are two essential histidines in each binding site. The advantages of using two modification reactions, ethoxyformylation and photooxidation, in the study of the functional role of histidines in proteins are demonstrated in this work.