Abstract：
Membrane proteins are essential components of biological membranes with key roles in cellular processes such as nutrient transport, cell communication, signaling, or energy conversion. Due to their crucial functions, membrane proteins and their complexes are often targets for therapeutic interventions. Expression and purification of membrane proteins are often a bottleneck to yield sufficient material for structural studies and further downstream characterization. Taking advantage of the Expi293 expression system for the production of eukaryotic proteins, we present a very efficient and fast protocol for the co-expression of a membrane complex. Here, we use transient transfection to co-express the membrane transporter PHT1 with its adaptor protein TASL. To allow the simultaneous screening of different proteins, constructs, or interaction partners, we make use of the Twin-Strep magnetic system. The protocol can be applied for small-scale screening of any membrane protein alone or co-expressed with interacting partners followed by large-scale production and purification of a potential membrane protein complex.
摘要：
膜蛋白是生物膜的重要组成部分，在细胞过程如营养运输中起关键作用。细胞通讯，信令,或能量转换。由于它们的关键功能，膜蛋白及其复合物通常是治疗干预的目标。膜蛋白的表达和纯化通常是产生用于结构研究和进一步下游表征的足够材料的瓶颈。利用Expi293表达系统生产真核蛋白质，我们提出了一种非常有效和快速的共表达膜复合物的方案。这里,我们使用瞬时转染共表达膜转运蛋白PHT1及其衔接蛋白TASL。为了允许同时筛选不同的蛋白质，constructs,或者互动伙伴，我们利用了Twin-Strep磁系统.该方案可应用于小规模筛选任何单独或与相互作用配偶共表达的膜蛋白，然后大规模生产和纯化潜在的膜蛋白复合物。
