Pandey Yogesh, Dondapati Srujan Kumar, Wüstenhagen Doreen, Kubick Stefan
Fraunhofer Institute for Cell Therapy and Immunology (IZI), Branch Bioanalytics and Bioprocesses (IZI-BB), Potsdam, Germany.
Institut für Biochemie und Biologie, University of Potsdam, Potsdam, OT Golm, Germany.
Adv Biochem Eng Biotechnol. 2023;186:103-120. doi: 10.1007/10_2023_228.
Cell-free protein synthesis (CFPS) has emerged as a powerful tool for the rapid synthesis and analysis of various structurally and functionally distinct proteins. These include 'difficult-to-express' membrane proteins such as large multipass ion channel receptors. Owing to their membrane localization, eukaryotic CFPS supplemented with endoplasmic reticulum (ER)-derived microsomal vesicles has proven to be an efficient system for the synthesis of functional membrane proteins. Here we demonstrate the applicability of the eukaryotic cell-free systems based on lysates from the mammalian Chinese Hamster Ovary (CHO) and insect Spodoptera frugiperda (Sf21) cells. We demonstrate the efficiency of the systems in the de novo cell-free synthesis of the human cardiac ion channels: ether-a-go-go potassium channel (hERG) K11.1 and the voltage-gated sodium channel hNa1.5.
无细胞蛋白质合成（CFPS）已成为一种强大的工具，可用于快速合成和分析各种结构和功能不同的蛋白质。这些蛋白质包括“难以表达”的膜蛋白，如大型多次跨膜离子通道受体。由于它们的膜定位，补充有内质网（ER）衍生微粒体囊泡的真核CFPS已被证明是合成功能性膜蛋白的有效系统。在这里，我们展示了基于哺乳动物中国仓鼠卵巢（CHO）细胞和昆虫草地贪夜蛾（Sf21）细胞裂解物的真核无细胞系统的适用性。我们展示了该系统在从头无细胞合成人类心脏离子通道：去极化激活钾通道（hERG）K11.1和电压门控钠通道hNa1.5方面的效率。
