Barouch W, Prasad K, Greene L E, Eisenberg E
Laboratory of Cell Biology, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland 20892.
J Biol Chem. 1994 Nov 18;269(46):28563-8.
In the presence of ATP, bovine brain hsp70 has been shown to remove clathrin from bovine brain clathrin-coated vesicles in a rapid stoichiometric initial burst followed by slow steady-state uncoating. In addition, it has been found recently that a 100-kDa cofactor is required for hsp70 to uncoat clathrin baskets prepared with the assembly protein AP-2. In this study the ATPase activity associated with uncoating was investigated, with baskets formed from clathrin and assembly proteins. Mixed assembly proteins or assembly protein AP-2 could not be used in ATPase studies because they activated the hsp70 ATPase activity even in the absence of clathrin. However, this was not the case with assembly protein AP180. A stoichiometric initial burst of ATP hydrolysis was found to accompany the initial burst of uncoating of AP180-clathrin baskets by hsp70, with 1 mol of hydrolyzed ATP/mol of released clathrin heavy chain. Furthermore, the presence of a 100-kDa cofactor was needed for both processes. These results suggest that an initial burst of uncoating occurs with all clathrin baskets, that an initial burst of ATP hydrolysis accompanies this initial burst of uncoating, and that a 100-kDa cofactor is required for both.
在ATP存在的情况下，已表明牛脑热休克蛋白70（hsp70）能迅速以化学计量的初始爆发形式从牛脑网格蛋白包被小泡中去除网格蛋白，随后是缓慢的稳态脱包被过程。此外，最近还发现，hsp70要使由装配蛋白AP-2制备的网格蛋白篮脱包被，需要一种100 kDa的辅因子。在本研究中，对与脱包被相关的ATP酶活性进行了研究，所用的网格蛋白篮由网格蛋白和装配蛋白形成。混合装配蛋白或装配蛋白AP-2不能用于ATP酶研究，因为它们即使在没有网格蛋白的情况下也能激活hsp70的ATP酶活性。然而，装配蛋白AP180的情况并非如此。发现hsp70使AP180-网格蛋白篮脱包被的初始爆发伴随着化学计量的ATP水解初始爆发，即每摩尔释放的网格蛋白重链有1摩尔水解的ATP。此外，这两个过程都需要100 kDa辅因子的存在。这些结果表明，所有网格蛋白篮都会发生初始脱包被爆发，ATP水解的初始爆发伴随着这种初始脱包被爆发，并且这两个过程都需要100 kDa辅因子。
