Galante Y M, Wong S Y, Hatefi Y
Biochemistry. 1981 Apr 28;20(9):2671-8. doi: 10.1021/bi00512a048.
Mitochondrial ATPase inhibitor protein (IF1) reacts reversibly with complex V and inhibits up to 90% of its ATPase activity. Both the rate and extent of inhibition are pH and temperature dependent and increase as the pH is lowered from pH 8 tp 6.7 (the lowest pH examined) or as the temperature is increased from 4 to 36 degrees C. Nucleotide triphosphates plus Mg2+ ions are required for inhibition of complex V ATPase activity by IF1. In the presence of Mg2+ ions, the effectiveness order of nucleotides is ATP greater than ITP greater than GTP greater than UTP. Highly purified complex V, which requires added phospholipids for expressing ATPase and ATP-Pi exchange activities, cannot be inhibited by IF1 plust ATP-Mg2+ unless phospholipids are also added. This indicates that the active state of the enzyme is necessary for the IF1 effect to be manifested, because F1-ATPase, which does not contain nor require phospholipids for catalyzing ATP hydrolysis, can be inhibited by IF1 plus ATP-Mg2+ in the absence of added phospholipids. The IF1-inhibited complex V, but not IF1-inhibited F1-ATPase, can be reactivated by incubation at pH greater than 7.0 in the absence of ATP-Mg2+. The reactivation rate is pH dependent and is influenced by temperature and enzyme concentration. Complex V preparations contain small and variable amounts of IF1. This endogenous IF1 behaves the same as added IF1 with respect to conditions described above for inhibition and reactivation and can result in 25-50% inhibition in different complex V preparations. However, complex V lacking endogenous IF1 can be reconstituted from F0, F1, oligomycin sensitivity conferring protein, and phospholipids. Inhibition of this reconstituted preparation in the presence of ATP-Mg2+ depends entirely on addition of IF1. In general, the ATP-Pi exchange activity of complex V is more sensitive to the chemical inhibitors of F1-AtPase tha its ATPase activity. This is not so, however, for IF1. Under conditions that IF1 caused approximately 75% inhibition of ATPase activity of complex V, no more than 10% of the ATP-Pi exchange activity was inhibited.
线粒体ATP酶抑制蛋白（IF1）与复合体V发生可逆反应，可抑制其高达90%的ATP酶活性。抑制的速率和程度均依赖于pH值和温度，当pH值从8降至6.7（所检测的最低pH值）或温度从4℃升至36℃时，抑制作用增强。IF1抑制复合体V的ATP酶活性需要三磷酸核苷酸和Mg2+离子。在Mg2+离子存在的情况下，核苷酸的有效作用顺序为ATP＞ITP＞GTP＞UTP。高度纯化的复合体V需要添加磷脂才能表现出ATP酶和ATP - Pi交换活性，除非也添加磷脂，否则IF1加ATP - Mg2+不能抑制它。这表明酶的活性状态是IF1发挥作用所必需的，因为在不添加磷脂的情况下，不含也不需要磷脂来催化ATP水解的F1 - ATP酶可被IF1加ATP - Mg2+抑制。IF1抑制的复合体V，但不包括IF1抑制的F1 - ATP酶，在不存在ATP - Mg2+的情况下，于pH值大于7.0孵育可被重新激活。重新激活速率依赖于pH值，并受温度和酶浓度影响。复合体V制剂含有少量且数量可变的IF1。就上述抑制和重新激活的条件而言，这种内源性IF1的行为与添加的IF1相同，并且在不同的复合体V制剂中可导致25% - 50%的抑制。然而，缺乏内源性IF1的复合体V可由F0、F1、赋予寡霉素敏感性的蛋白和磷脂重新组装而成。在ATP - Mg2+存在的情况下，这种重新组装制剂的抑制完全取决于IF1的添加。一般来说，复合体V的ATP - Pi交换活性比其ATP酶活性对F1 - ATP酶的化学抑制剂更敏感。然而，IF1并非如此。在IF1导致复合体V的ATP酶活性约75%被抑制的条件下，ATP - Pi交换活性的抑制不超过10%。
