Mizrachi Dario, Chen Yujie, Liu Jiayan, Peng Hwei-Ming, Ke Ailong, Pollack Lois, Turner Raymond J, Auchus Richard J, DeLisa Matthew P
School of Chemical and Biomolecular Engineering, Cornell University, Ithaca, New York 14853, USA.
School of Applied and Engineering Physics, Cornell University, Ithaca, New York 14853, USA.
Nat Commun. 2015 Apr 8;6:6826. doi: 10.1038/ncomms7826.
Integral membrane proteins (IMPs) play crucial roles in all cells and represent attractive pharmacological targets. However, functional and structural studies of IMPs are hindered by their hydrophobic nature and the fact that they are generally unstable following extraction from their native membrane environment using detergents. Here we devise a general strategy for in vivo solubilization of IMPs in structurally relevant conformations without the need for detergents or mutations to the IMP itself, as an alternative to extraction and in vitro solubilization. This technique, called SIMPLEx (solubilization of IMPs with high levels of expression), allows the direct expression of soluble products in living cells by simply fusing an IMP target with truncated apolipoprotein A-I, which serves as an amphipathic proteic 'shield' that sequesters the IMP from water and promotes its solubilization.
整合膜蛋白（IMPs）在所有细胞中都发挥着关键作用，是颇具吸引力的药理学靶点。然而，IMPs的功能和结构研究受到其疏水性的阻碍，而且使用去污剂从其天然膜环境中提取后，它们通常不稳定。在此，我们设计了一种通用策略，可在不使用去污剂或对IMPs本身进行突变的情况下，在体内以结构相关的构象溶解IMPs，作为提取和体外溶解的替代方法。这种技术称为SIMPLEx（高水平表达溶解IMPs），通过简单地将IMPs靶标与截短的载脂蛋白A-I融合，使可溶性产物在活细胞中直接表达，截短的载脂蛋白A-I作为一种两亲性蛋白质“护盾”，将IMPs与水隔离并促进其溶解。
