Galmiche J M, Girault G, Berger G, Jacquot J P, Miginiac-Maslow M, Wollman E
Service de Biophysique, CEN Saclay, Gif-sur-Yvette, France.
Biochimie. 1990 Jan;72(1):25-32. doi: 10.1016/0300-9084(90)90169-h.
ATPase activity of the coupling factor 1, CF1, isolated from spinach chloroplasts, was enhanced by reduction with dithiothreitol. Reduced thioredoxins from spinach chloroplasts, Escherichia coli and human lymphocytes replaced dithiothreitol as reductant and activator of the ATPase. CF1 must be in an oxidized activated state to be further activated by reduced thioredoxin. This state was obtained either by heating CF1 or removing the inhibitory intrinsic epsilon subunit from CF1. Efficiency and primary structure of the different thioredoxins were compared. The progressive addition of KCl during ATPase activation by reduced thioredoxin increases then decreases this process. We proposed that three basic amino acids corresponding to arginine 73 and lysines 82 and 96 in Escherichia coli thioredoxin play an important role in the anchorage of the thioredoxin to the negatively charged surface of the CF1 and are involved in the dual effect of KCl. The variations in the screening effect of the negative charges of the CF1 surface by K+ ions can indeed explain the changes in the anchorage of these 3 basic amino acids with concomitant variation in ATPase activity. Human thioredoxin must be 10 times more concentrated than Escherichia coli or spinach chloroplast thioredoxin to exhibit the same activation effect on the ATPase. This fact was related to the properties of a sequence equivalent to the part from amino acid 59 to 72 in Escherichia coli thioredoxin. This part which joins the two lobes of the thioredoxin is more hydrophilic and more negatively charged in human thioredoxin than in Escherichia coli or spinach chloroplast thioredoxin. Although ATPase activation was obtained at a very low concentration of the reduced spinach chloroplast thioredoxin, the thioredoxin formed only a loose complex with CF1.
从菠菜叶绿体中分离出的偶联因子1（CF1）的ATP酶活性，通过二硫苏糖醇还原得到增强。来自菠菜叶绿体、大肠杆菌和人淋巴细胞的还原型硫氧还蛋白替代二硫苏糖醇作为ATP酶的还原剂和激活剂。CF1必须处于氧化激活状态才能被还原型硫氧还蛋白进一步激活。这种状态可以通过加热CF1或从CF1中去除抑制性的内在ε亚基来获得。比较了不同硫氧还蛋白的效率和一级结构。在还原型硫氧还蛋白激活ATP酶的过程中逐步添加KCl，该过程先增加后减少。我们提出，大肠杆菌硫氧还蛋白中与精氨酸73以及赖氨酸82和96相对应的三个碱性氨基酸，在硫氧还蛋白与带负电荷的CF1表面的锚定中起重要作用，并参与KCl的双重作用。K⁺离子对CF1表面负电荷的屏蔽作用的变化确实可以解释这三个碱性氨基酸锚定的变化以及ATP酶活性的相应变化。人硫氧还蛋白的浓度必须比大肠杆菌或菠菜叶绿体硫氧还蛋白高10倍才能对ATP酶表现出相同的激活作用。这一事实与大肠杆菌硫氧还蛋白中相当于氨基酸59至72部分的序列特性有关。在人硫氧还蛋白中，连接硫氧还蛋白两个叶的这部分比大肠杆菌或菠菜叶绿体硫氧还蛋白更亲水且带更多负电荷。尽管在极低浓度的还原型菠菜叶绿体硫氧还蛋白下即可实现ATP酶的激活，但硫氧还蛋白与CF1仅形成一种松散的复合物。
