Armon T, Ganoth D, Hershko A
Unit of Biochemistry, Faculty of Medicine, Technion-Israel Institute of Technology, Haifa.
J Biol Chem. 1990 Dec 5;265(34):20723-6.
In the ubiquitin pathway for intracellular protein breakdown, proteins ligated to ubiquitin are degraded by a large (26 S) ATP-dependent protease complex. It was found previously that the 26 S complex is assembled from three different enzyme components by a process that requires MgATP. In addition, MgATP is also required for the continued action of the 26 S complex in the breakdown of ubiquitin-protein conjugates. In the present study we have tried to gain some insight into the mode of action of ATP by following ATP hydrolysis by the 26 S complex and its three components. It was found that none of the three unassembled components had significant ATPase activity, but such activity appeared following their entry into the 26 S complex. The presence of all three components and of MgATP was required for the formation of complex-associated ATPase activity. GTP and UTP cannot replace ATP for complex assembly, but these nucleotides can substitute for ATP in the stimulation of the conjugate-degrading activity of the 26 S complex. Unlabeled GTP and UTP inhibit the hydrolysis of [gamma-32P] ATP by complex-associated ATPase, indicating that this activity is related to the latter site of ATP action in this system.
在细胞内蛋白质降解的泛素途径中，与泛素连接的蛋白质被一种大型（26S）依赖ATP的蛋白酶复合体降解。先前发现，26S复合体是由三种不同的酶组分通过一个需要MgATP的过程组装而成。此外，MgATP对于26S复合体在泛素 - 蛋白质缀合物降解中的持续作用也是必需的。在本研究中，我们试图通过跟踪26S复合体及其三个组分的ATP水解情况，来深入了解ATP的作用方式。结果发现，三种未组装的组分均无显著的ATP酶活性，但它们进入26S复合体后会出现这种活性。形成与复合体相关的ATP酶活性需要所有三种组分和MgATP的存在。GTP和UTP不能替代ATP进行复合体组装，但这些核苷酸可以在刺激26S复合体的缀合物降解活性方面替代ATP。未标记的GTP和UTP会抑制与复合体相关的ATP酶对[γ - 32P]ATP的水解，这表明该活性与该系统中ATP作用的后一个位点有关。
