Plemper R K, Deak P M, Otto R T, Wolf D H
Institut für Biochemie, Universität Stuttgart, Germany.
FEBS Lett. 1999 Jan 29;443(3):241-5. doi: 10.1016/s0014-5793(98)01724-4.
Misfolded or unassembled secretory proteins are retained in the endoplasmic reticulum (ER) and subsequently degraded by the cytosolic ubiquitin-proteasome system. This requires their retrograde transport from the ER lumen into the cytosol, which is mediated by the Sec61 translocon. It had remained a mystery whether ER-localised soluble proteins are at all capable of re-entering the Sec61 channel de novo or whether a permanent contact of the imported protein with the translocon is a prerequisite for retrograde transport. In this study we analysed two new variants of the mutated yeast carboxypeptidase yscY, CPY*: a carboxy-terminal fusion protein of CPY* and pig liver esterase and a CPY* species carrying an additional glycosylation site at its carboxy-terminus. With these constructs it can be demonstrated that the newly synthesised CPY* chain is not retained in the translocation channel but reaches its ER lumenal side completely. Our data indicate that the Sec61 channel provides the essential pore for protein transport through the ER membrane in either direction; persistent contact with the translocon after import seems not to be required for retrograde transport.
错误折叠或未组装的分泌蛋白会滞留在内质网（ER）中，随后被胞质泛素 - 蛋白酶体系统降解。这需要它们从内质网腔逆向转运到胞质溶胶中，此过程由Sec61转运体介导。内质网定位的可溶性蛋白是否能够重新从头进入Sec61通道，或者导入的蛋白与转运体的永久接触是否是逆向转运的先决条件，一直是个谜。在本研究中，我们分析了突变酵母羧肽酶yscY（CPY*）的两个新变体：CPY与猪肝酯酶的羧基末端融合蛋白以及在其羧基末端带有额外糖基化位点的CPY物种。通过这些构建体可以证明，新合成的CPY*链不会保留在转运通道中，而是完全到达其内质网腔侧。我们的数据表明，Sec61通道为蛋白质双向穿过内质网膜提供了必要的孔道；导入后与转运体的持续接触似乎不是逆向转运所必需的。
