Manley Suliana, Horton Margaret R, Lecszynski Szymon, Gast Alice P
Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, USA.
Biophys J. 2008 Sep;95(5):2301-7. doi: 10.1529/biophysj.107.124024. Epub 2008 May 23.
Heterogeneities in cell membranes due to the ordering of lipids and proteins are thought to play an important role in enabling protein and lipid trafficking throughout the secretory pathway and in maintaining cell polarization. Protein-coated vesicles provide a major mechanism for intracellular transport of select cargo, which may be sorted into lipid microdomains; however, the mechanisms and physical constraints for lipid sorting by protein coats are relatively unexplored. We studied the influence of membrane-tethered protein coats on the sorting, morphology, and phase behavior of liquid-ordered lipid domains in a model system of giant unilamellar vesicles composed of dioleoylphosphatidylcholine, sphingomyelin, and cholesterol. We created protein-coated membranes by forming giant unilamellar vesicles containing a small amount of biotinylated lipid, thereby creating binding sites for streptavidin and avidin proteins in solution. We found that individual tethered proteins colocalize with the liquid-disordered phase, whereas ordered protein domains on the membrane surface colocalize with the liquid-ordered phase. These observations may be explained by considering the thermodynamics of this coupled system, which maximizes its entropy by cosegregating ordered protein and lipid domains. In addition, protein ordering inhibits lipid domain rearrangement and modifies the morphology and miscibility transition temperature of the membrane, most dramatically near the critical point in the membrane phase diagram. This observation suggests that liquid-ordered domains are stabilized by contact with ordered protein domains; it also hints at an approach to the stabilization of lipid microdomains by cross-linked protein clusters or ordered protein coats.
由于脂质和蛋白质的有序排列导致的细胞膜异质性，被认为在促进蛋白质和脂质在整个分泌途径中的运输以及维持细胞极化方面发挥着重要作用。蛋白质包被的囊泡为特定货物的细胞内运输提供了一种主要机制，这些货物可能被分选到脂质微区中；然而，蛋白质包被对脂质分选的机制和物理限制相对尚未被探索。我们在由二油酰磷脂酰胆碱、鞘磷脂和胆固醇组成的巨型单层囊泡模型系统中，研究了膜 tethered 蛋白包被对液相有序脂质域的分选、形态和相行为的影响。我们通过形成含有少量生物素化脂质的巨型单层囊泡来创建蛋白质包被的膜，从而在溶液中为链霉亲和素和抗生物素蛋白创造结合位点。我们发现单个 tethered 蛋白与液相无序相共定位，而膜表面上的有序蛋白域与液相有序相共定位。通过考虑这个耦合系统的热力学可以解释这些观察结果，该系统通过将有序的蛋白质和脂质域共分离来最大化其熵。此外，蛋白质有序化抑制脂质域重排，并改变膜在膜相图临界点附近最显著的形态和混溶转变温度。这一观察结果表明，液相有序域通过与有序蛋白域接触而稳定；它还暗示了一种通过交联蛋白簇或有序蛋白包被来稳定脂质微区的方法。
