Reinvestigation of the requirement of cytosolic ATP for mitochondrial protein import
Reinvestigation of the requirement of cytosolic ATP for mitochondrial protein import
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DOI：
10.1074/jbc.m401291200
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发表时间：
2004-05-07
影响因子：
4.8
通讯作者：
Endo, T
中科院分区：
文献类型：
--
作者：
Asai, T;Takahashi, T;Endo, T
蛋白质进入线粒体需要线粒体内外ATP水解的能量。虽然线粒体基质中的ATP在线粒体蛋白输入中的作用已被广泛研究，但线粒体外的ATP（外部ATP）的作用仍然很少被表征。在这里，我们开发了一种方案，用于耗尽外部ATP，而不显着降低进口能力的前体蛋白在体外合成的网织红细胞裂解物。我们测试了外部ATP对各种前体蛋白进入分离的酵母线粒体的影响。我们发现，外部ATP是必需的线粒体前体蛋白的进口能力的维护，但一旦它们结合到线粒体，随后的易位的前序列包含的蛋白质，但不是ADP/ATP载体，独立于外部ATP进行。由于细胞溶质Hsp 70的耗竭导致线粒体前体蛋白的输入能力降低，因此细胞溶质Hsp 70可能利用外部ATP。相比之下，ADP/ATP载体需要外部ATP以有效地输入到线粒体中，即使在与线粒体结合后，这种情况仅部分归因于胞质Hsp 70。
Protein import into mitochondria requires the energy of ATP hydrolysis inside and/or outside mitochondria. Although the role of ATP in the mitochondrial matrix in mitochondrial protein import has been extensively studied, the role of ATP outside mitochondria (external ATP) remains only poorly characterized. Here we developed a protocol for depletion of external ATP without significantly reducing the import competence of precursor proteins synthesized in vitro with reticulocyte lysate. We tested the effects of external ATP on the import of various precursor proteins into isolated yeast mitochondria. We found that external ATP is required for maintenance of the import competence of mitochondrial precursor proteins but that, once they bind to mitochondria, the subsequent translocation of presequence-containing proteins, but not the ADP/ATP carrier, proceeds independently of external ATP. Because depletion of cytosolic Hsp70 led to a decrease in the import competence of mitochondrial precursor proteins, external ATP is likely utilized by cytosolic Hsp70. In contrast, the ADP/ATP carrier requires external ATP for efficient import into mitochondria even after binding to mitochondria, a situation that is only partly attributed to cytosolic Hsp70.