A novel yeast expression system for the overproduction of quality-controlled membrane proteins
A novel yeast expression system for the overproduction of quality-controlled membrane proteins
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DOI：
10.1016/s0014-5793(03)00952-9
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发表时间：
2003-10-09
期刊：
影响因子：
3.5
通讯作者：
Oesterhelt, D
中科院分区：
文献类型：
--
作者：
Griffith, DA;Delipala, C;Oesterhelt, D
细胞蛋白质折叠能力的饱和和非活性不完全折叠蛋白的积累往往伴随着膜蛋白（MPs）的过度表达，这对膜蛋白在功能形式下的有效纯化和结构研究构成了障碍。我们提出了一种优化酿酒酵母NIP功能表达的新策略。该方法利用未折叠蛋白反应（UPR）途径，这是一种应激信号机制，可感知内质网中未折叠蛋白的积累。我们证明，高水平的UPR诱导NIP的表达反映了该蛋白的功能表达受损。调节蛋白的表达水平以避免或减少UPR诱导导致其功能表达增加。因此，UPR状态可以作为MP表达受损程度的代理变量，甚至可以在不了解蛋白质生物学功能的情况下适用。(C) 2003年欧洲生化学会联合会。Elsevier B.V.版权所有。
Saturation of the cell's protein folding capacity and accumulation of inactive incompletely folded protein often accompanying the overexpression of membrane proteins (MPs) presents an obstacle to their efficient purification in a functional form for structural studies. We present a novel strategy for optimization of functional NIP expression in Saccharomyces cerevisiae. This approach exploits the unfolded protein response (UPR) pathway, a stress signaling mechanism that senses the accumulation of unfolded proteins in the endoplasmic reticulum. We demonstrate that a high level of UPR induction upon expression of a NIP reflects impaired functional expression of that protein. Tuning the expression level of the protein so as to avoid or minimize UPR induction results in its increased functional expression. UPR status can therefore serve as a proxy variable for the extent of impaired expression of a MP that may even be applicable in the absence of knowledge of the protein's biological function. (C) 2003 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.