Department of Chemistry and Biochemistry, University of Denver, Denver, CO 80208, USA.
Soft Matter. 2014 Mar 28;10(12):2016-23. doi: 10.1039/c3sm52522h.
Cellular membranes contain a variety of shapes that likely act as motifs for sorting lipids and proteins. To understand the sorting that takes place within cells, a continuous, fluid bilayer with regions of membrane curvature was designed and characterized using confocal fluorescence and total internal reflection fluorescence microscopy techniques. A supported lipid bilayer was formed over fluorescently labelled nanoparticles deposited on a glass surface. The lipid composition and membrane shape are separately controlled and the nanoparticle dimensions (d = 40-200 nm) determine the extent of curvature. The bulk membrane is fluid as demonstrated by fluorescence recovery after photobleaching (FRAP) using dye labelled lipids. In bilayers that contain fluorescently labelled, single-tailed lipids, accumulation is observed at regions of curvature, yet the molecules retain fluidity. Using single particle imaging methods, lipids are observed to visit regions of curvature and exchange with the surrounding flat membrane. The nanoparticle patterned substrate described here allows for quantitative measurement of the transient interactions between fluorescently labelled biomolecules and regions of membrane curvature.
细胞膜含有多种形状，这些形状可能充当脂质和蛋白质分选的模式。为了理解细胞内发生的分选，设计并使用共聚焦荧光和全内反射荧光显微镜技术对具有膜曲率区域的连续、流体双层膜进行了表征。在玻璃表面上沉积荧光标记纳米颗粒的支持脂质双层。分别控制脂质组成和膜形状，纳米颗粒的尺寸（d=40-200nm）决定曲率的程度。通过使用染料标记的脂质进行荧光漂白后荧光恢复（FRAP）实验，证明了大块膜是流体的。在含有荧光标记的单尾脂质的双层膜中，在曲率区域观察到了积累，但这些分子仍保持流动性。使用单颗粒成像方法，观察到脂质能够访问曲率区域并与周围的平面膜进行交换。这里描述的纳米颗粒图案化基底允许对荧光标记的生物分子与膜曲率区域之间的瞬时相互作用进行定量测量。
