Measuring the Energetics of Membrane Protein Dimerization in Mammalian Membranes
Measuring the Energetics of Membrane Protein Dimerization in Mammalian Membranes
复制标题
DOI：
10.1021/ja910692u
复制
发表时间：
2010-03-17
影响因子：
15
通讯作者：
Hristova, Kalina
中科院分区：
文献类型：
--
作者：
Chen, Lirong;Novicky, Lawrence;Hristova, Kalina
到目前为止，对膜中横向蛋白质相互作用的定量研究一直局限于脂泡或细菌膜中的多肽或简化的蛋白质结构。在这里，我们展示了如何在哺乳动物质膜来源的囊泡中测量膜蛋白二聚的自由能。测量在单个囊泡中进行，利用定量成像FRET(QI-FRET)方法。这些实验是以循序渐进的方式描述的。该蛋白的特征是血糖蛋白A的跨膜区，它是研究最广泛的膜蛋白，已知在疏水环境中形成同源二聚体。结果表明，细胞膜中的分子拥挤对膜蛋白相互作用的强度有很大的影响。
Thus far, quantitative studies of lateral protein interactions in membranes have been restricted peptides or simplified protein constructs in lipid vesicles or bacterial membranes. Here we show how free energies of membrane protein dimerization can be measured in mammalian plasma membrane-derived vesicles. The measurements, performed in single vesicles, utilize the quantitative imaging FRET (QI-FRET) method. The experiments are described in a step-by-step protocol. The protein characterized is the transmembrane domain of glycophorin A, the most extensively studied membrane protein, known to form homodimers in hydrophobic environments. The results suggest that molecular crowding in cellular membranes has a dramatic effect on the strength of membrane protein interactions.