Mayer A, Lill R, Neupert W
Institut für Physiologische Chemie, Universität München, Federal Republic of Germany.
J Cell Biol. 1993 Jun;121(6):1233-43. doi: 10.1083/jcb.121.6.1233.
Nuclear-encoded proteins destined for mitochondria must cross the outer or both outer and inner membranes to reach their final sub-mitochondrial locations. While the inner membrane can translocate preproteins by itself, it is not known whether the outer membrane also contains an endogenous protein translocation activity which can function independently of the inner membrane. To selectively study the protein transport into and across the outer membrane of Neurospora crassa mitochondria, outer membrane vesicles were isolated which were sealed, in a right-side-out orientation, and virtually free of inner membranes. The vesicles were functional in the insertion and assembly of various outer membrane proteins such as porin, MOM19, and MOM22. Like with intact mitochondria, import into isolated outer membranes was dependent on protease-sensitive surface receptors and led to correct folding and membrane integration. The vesicles were also capable of importing a peripheral component of the inner membrane, cytochrome c heme lyase (CCHL), in a receptor-dependent fashion. Thus, the protein translocation machinery of the outer mitochondrial membrane can function as an independent entity which recognizes, inserts, and translocates mitochondrial preproteins of the outer membrane and the intermembrane space. In contrast, proteins which have to be translocated into or across the inner membrane were only specifically bound to the vesicles, but not imported. This suggests that transport of such proteins involves the participation of components of the intermembrane space and/or the inner membrane, and that in these cases the outer membrane translocation machinery has to act in concert with that of the inner membrane.
destined for mitochondria must cross the outer or both outer and inner membranes to reach their final sub-mitochondrial locations. While the inner membrane can translocate preproteins by itself, it is not known whether the outer membrane also contains an endogenous protein translocation activity which can function independently of the inner membrane. To selectively study the protein transport into and across the outer membrane of Neurospora crassa mitochondria, outer membrane vesicles were isolated which were sealed, in a right-side-out orientation, and virtually free of inner membranes. The vesicles were functional in the insertion and assembly of various outer membrane proteins such as porin, MOM19, and MOM22. Like with intact mitochondria, import into isolated outer membranes was dependent on protease-sensitive surface receptors and led to correct folding and membrane integration. The vesicles were also capable of importing a peripheral component of the inner membrane, cytochrome c heme lyase (CCHL), in a receptor-dependent fashion. Thus, the protein translocation machinery of the outer mitochondrial membrane can function as an independent entity which recognizes, inserts, and translocates mitochondrial preproteins of the outer membrane and the intermembrane space. In contrast, proteins which have to be translocated into or across the inner membrane were only specifically bound to the vesicles, but not imported. This suggests that transport of such proteins involves the participation of components of the intermembrane space and/or the inner membrane, and that in these cases the outer membrane translocation machinery has to act in concert with that of the inner membrane.
定位于线粒体的核编码蛋白必须穿过外膜或同时穿过外膜和内膜，才能到达其最终的亚线粒体位置。虽然内膜自身能够转运前体蛋白，但尚不清楚外膜是否也含有一种可独立于内膜发挥作用的内源性蛋白转运活性。为了选择性地研究蛋白质进入粗糙脉孢菌线粒体的外膜并穿过外膜的过程，分离出了外膜囊泡。这些囊泡是外翻定向密封的，且几乎不含内膜。这些囊泡在各种外膜蛋白如孔蛋白、MOM19和MOM22 的插入和组装过程中发挥作用。与完整线粒体一样，蛋白质导入分离的外膜依赖于蛋白酶敏感的表面受体，并导致正确折叠和膜整合。这些囊泡还能够以受体依赖的方式导入内膜的外周成分——细胞色素c血红素裂解酶（CCHL）。因此，线粒体外膜的蛋白质转运机制可以作为一个独立的实体发挥作用，识别、插入并转运外膜和膜间隙的线粒体前体蛋白。相比之下，必须转运到内膜内或穿过内膜的蛋白质仅特异性地结合到囊泡上，但未被导入。这表明此类蛋白质的转运涉及膜间隙和/或内膜成分的参与，并且在这些情况下，外膜转运机制必须与内膜转运机制协同作用。
