Membrane Biochemistry and Biophysics, Bijvoet Center for Biomolecular Research, Institute of Biomembranes, Utrecht University, Padualaan 8, 3584 CH Utrecht, The Netherlands.
Cell Biology, Department of Biology, Faculty of Science, Utrecht University, Padualaan 8, 3584 CH Utrecht, The Netherlands.
Biochim Biophys Acta Biomembr. 2017 Nov;1859(11):2155-2160. doi: 10.1016/j.bbamem.2017.08.010. Epub 2017 Aug 25.
Extracting membrane proteins from biological membranes by styrene-maleic acid copolymers (SMAs) in the form of nanodiscs has developed into a powerful tool in membrane research. However, the mode of action of membrane (protein) solubilization in a cellular context is still poorly understood and potential specificity for cellular compartments has not been investigated. Here, we use fluorescence microscopy to visualize the process of SMA solubilization of human cells, exemplified by the immortalized human HeLa cell line. Using fluorescent protein fusion constructs that mark distinct subcellular compartments, we found that SMA solubilizes membranes in a concentration-dependent multi-stage process. While all major intracellular compartments were affected without a strong preference, plasma membrane solubilization was found to be generally slower than the solubilization of organelle membranes. Interestingly, some plasma membrane-localized proteins were more resistant against solubilization than others, which might be explained by their presence in specific membrane domains with differing properties. Our results support the general applicability of SMA for the isolation of membrane proteins from different types of (sub)cellular membranes.
通过苯乙烯-马来酸共聚物（SMAs）纳米盘的形式从生物膜中提取膜蛋白已成为膜研究中的有力工具。然而，在细胞环境中膜（蛋白）溶解的作用模式仍知之甚少，且尚未研究其对细胞区室的潜在特异性。在这里，我们使用荧光显微镜可视化 SMA 溶解人细胞的过程，以永生的人宫颈癌细胞系 HeLa 为例。使用标记不同亚细胞区室的荧光蛋白融合构建体，我们发现 SMA 以浓度依赖的多阶段过程溶解膜。虽然所有主要的细胞内区室都受到影响而没有强烈的偏好，但发现质膜的溶解通常比细胞器膜的溶解慢。有趣的是，一些定位于质膜的蛋白质比其他蛋白质更能抵抗溶解，这可能是由于它们存在于具有不同性质的特定膜域中。我们的结果支持 SMA 用于从不同类型的（亚）细胞膜中分离膜蛋白的一般适用性。
