Mapping the Cargo Protein Membrane Translocation Step into the PEX5 Cycling Pathway
Mapping the Cargo Protein Membrane Translocation Step into the PEX5 Cycling Pathway
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DOI：
10.1074/jbc.m109.032565
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发表时间：
2009-10-02
影响因子：
4.8
通讯作者：
Azevedo, Jorge E.
中科院分区：
文献类型：
--
作者：
Alencastre, Ines S.;Rodrigues, Tony A.;Azevedo, Jorge E.
新合成的过氧化物酶体基质蛋白通过过氧化物酶体循环受体 PEX5 靶向细胞器。在过去的几年中，利用以 PEX5 为中心的体外系统获得了有关该过程机制的宝贵数据。迄今为止收集的数据表明胞质 PEX5。货物蛋白复合物停靠在过氧化物酶体对接/易位机器上，PEX5 随后以不依赖于 ATP 的方式插入其中。然后，该 PEX5 物种在保守的半胱氨酸残基处被单泛素化，这是该途径下一步的强制性修饰，即泛素-PEX5 缀合物的 ATP 依赖性错位回到胞质溶胶中。最后，去除泛素部分，产生游离的 PEX5。尽管它很有用，但仍有许多未解决的机制方面无法用这种体外系统来解决，因此需要以货物蛋白为中心的视角。在这里，我们描述了一个强大的过氧化物酶体体外导入系统，它提供了这种视角。用它获得的数据表明，货物蛋白跨过氧化物酶体膜的易位，包括其释放到细胞器基质中，发生在PEX5泛素化之前。
Newly synthesized peroxisomal matrix proteins are targeted to the organelle by PEX5, the peroxisomal cycling receptor. Over the last few years, valuable data on the mechanism of this process have been obtained using a PEX5-centered in vitro system. The data gathered until now suggest that cytosolic PEX5. cargo protein complexes dock at the peroxisomal docking/translocation machinery, where PEX5 becomes subsequently inserted in an ATP-independent manner. This PEX5 species is then monoubiquitinated at a conserved cysteine residue, a mandatory modification for the next step of the pathway, the ATP-dependent dislocation of the ubiquitin-PEX5 conjugate back into the cytosol. Finally, the ubiquitin moiety is removed, yielding free PEX5. Despite its usefulness, there are many unsolved mechanistic aspects that cannot be addressed with this in vitro system and that call for a cargo protein-centered perspective instead. Here we describe a robust peroxisomal in vitro import system that provides this perspective. The data obtained with it suggest that translocation of a cargo protein across the peroxisomal membrane, including its release into the organelle matrix, occurs prior to PEX5 ubiquitination.