Center for Biomedical Engineering NE47-379, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Cambridge, MA 02139-4307, USA.
Proc Natl Acad Sci U S A. 2011 May 31;108(22):9049-54. doi: 10.1073/pnas.1018185108. Epub 2011 May 11.
Two major bottlenecks in elucidating the structure and function of membrane proteins are the difficulty of producing large quantities of functional receptors, and stabilizing them for a sufficient period of time. Selecting the right surfactant is thus crucial. Here we report using peptide surfactants in commercial Escherichia coli cell-free systems to rapidly produce milligram quantities of soluble G protein-coupled receptors (GPCRs). These include the human formyl peptide receptor, human trace amine-associated receptor, and two olfactory receptors. The GPCRs expressed in the presence of the peptide surfactants were soluble and had α-helical secondary structures, suggesting that they were properly folded. Microscale thermophoresis measurements showed that one olfactory receptor expressed using peptide surfactants bound its known ligand heptanal (molecular weight 114.18). These short and simple peptide surfactants may be able to facilitate the rapid production of GPCRs, or even other membrane proteins, for structure and function studies.
阐明膜蛋白结构和功能的两个主要瓶颈是大量生产功能受体的困难，以及为其提供足够长时间的稳定性。因此，选择合适的表面活性剂至关重要。在这里，我们报告了在商业大肠杆菌无细胞系统中使用肽表面活性剂来快速生产毫克级可溶性 G 蛋白偶联受体 (GPCR)。其中包括人源甲酰肽受体、人源痕迹胺相关受体和两种嗅觉受体。在肽表面活性剂存在下表达的 GPCR 是可溶性的，具有α-螺旋二级结构，表明它们正确折叠。微量热泳动测量表明，一种使用肽表面活性剂表达的嗅觉受体与其已知配体庚醛（分子量 114.18）结合。这些短而简单的肽表面活性剂可能能够促进 GPCR 甚至其他膜蛋白的快速生产，以进行结构和功能研究。
