Kübrich M, Rassow J, Voos W, Pfanner N, Hönlinger A
Institut für Biochemie und Molekularbiologie, Universität Freiburg, Hermann-Herder-Strabetae 7, D-79104 Freiburg, Germany.
J Biol Chem. 1998 Jun 26;273(26):16374-81. doi: 10.1074/jbc.273.26.16374.
The ADP/ATP carrier (AAC) of the mitochondrial inner membrane is synthesized in the cytosol without a cleavable presequence. The preprotein preferentially binds to the mitochondrial surface receptor Tom70 and joins the import pathway of presequence-carrying preproteins at the cis side of the outer membrane. Little is known about the translocation of the AAC across the outer membrane and where its import route separates from that of cleavable preproteins. Here we have characterized a translocation intermediate of AAC during transfer across the outer membrane. The major portion of the preprotein is exposed to the intermembrane space, while a short segment is still accessible to externally added protease. This intermediate can be quantitatively chased to the fully imported form in the inner membrane. Its accumulation depends on Tom7, but not on the intermembrane space domain of Tom22 in contrast to cleavable preproteins. Moreover, opening of the intermembrane space inhibits the import of AAC, but not that of cleavable preproteins into mitoplasts. We conclude that the import route of AAC diverges from the general import pathway of cleavable preproteins already at the trans side of the outer membrane.
线粒体内膜的ADP/ATP载体（AAC）在胞质溶胶中合成，没有可裂解的前导序列。前体蛋白优先结合到线粒体表面受体Tom70，并在外膜顺式侧加入携带前导序列的前体蛋白的导入途径。关于AAC穿过外膜的转运以及其导入途径与可裂解前体蛋白的导入途径在何处分离，人们知之甚少。在这里，我们表征了AAC在外膜转移过程中的转运中间体。前体蛋白的大部分暴露于膜间隙，而一小段仍可被外部添加的蛋白酶作用。这种中间体可以定量地追踪到内膜中的完全导入形式。其积累依赖于Tom7，但与可裂解前体蛋白不同，不依赖于Tom22的膜间隙结构域。此外，膜间隙的开放抑制了AAC的导入，但不抑制可裂解前体蛋白导入线粒体。我们得出结论，AAC的导入途径在外膜反式侧就已经与可裂解前体蛋白的一般导入途径不同。
