以螢光顯微鏡研究二元磷脂質人造膜及固醇加入後的效應

Fluorescence microscopy study of binary phospholipid membranes and the effect of sterol

廖國惠(Kou-Hwei Liao)

指導教授 ： 薛雅薇

國立中央大學/理學院/物理學系/碩士(2008年)

摘要

生物細胞膜是由脂質分子與膜蛋白所構成。許多實驗證據顯示不同種類的脂質分子混合後會有區塊(domains)的產生，而富含膽固醇(cholesterol)與抱合脂質(sphigolipid，屬於飽和脂質類)的脂質浮排(lipid raft)，被認為與細胞膜功能展現有重要的關聯。我們利用螢光顯微技術，佐以螢光分子NBD-PC偏好存在較無序區塊的特性，來研究含不飽和脂質及飽和脂質的人造膜系統(model membranes)，並探討固醇(sterol)加入的效應。研究發現，含不飽和脂質及飽和脂質(如DOPC/DPPC、POPC/DPPC)在低溫時會有亮暗區塊的產生。螢光亮區所顯示的應是液晶無序態(ld phase)的脂質分子，而暗區為處於膠態(so phase)的脂質。而在加入固醇後，原本的亮暗區塊會變的邊緣清晰、亮暗分明，此時螢光亮區代表的應是液晶無序態(ld phase)脂質分子，而暗區則是處於液晶有序態(lo phase)的脂質。而隨著溫度升高，處於無序的螢光亮區比例也會隨之增加，人造膜愈趨向無序。我們並發現脂質的碳氫鏈的不飽和度會影響本研究中的人造膜的有序程度。當不飽和碳氫鏈的數目增加時，人造膜會的有序程度會隨之降低。相較於含有膽固醇的人造膜，含有麥角固醇(ergosterol)的人造膜的暗區比例大幅減少，亦即，較無序。由此可知，固醇結構上的微小差異對人造膜的性質影響甚大。

關鍵字

細胞膜 ； 固醇 ； 螢光顯微鏡

並列摘要

Biological membranes are composed of lipids and proteins. Many evidences suggest that lipid domains coexist in biological membranes of living cells. The domains (lipid rafts) enriched in cholesterol and sphigolipid have been implicated to play important roles in a wide range of biological processes. In this work, we study the properties of model membranes composed of unsaturated and saturated lipid, and the effect of sterol using fluorescence microscopy. The fluorescent probe NBD-PC, preferentially located in a more disordered environment, is used in this experiment. We found that the bright and dark phases coexist in model membranes containing unsaturated and saturated lipids such as DOPC/DPPC and POPC/DPPC. The bright and dark phases correspond to where the lipids are in the ld and so phases, respectively. Addition of sterol to unsaturated lipid/saturated lipid membranes results in a more well-defined bright/dark boundary. In these sterol-containing membranes, the bright and dark phases correspond to where the lipids are in the ld and lo phases, respectively. The fraction of bright phase increases with temperature, indicating that membranes become more disordered as temperature is raised. We also found that the degree of lipid saturation affects the membranes order: the order of membranes decreases as the number of unsaturated chain increases. Replacing cholesterol with ergosterol substantially reduces the dark phase in unsaturated lipid/saturated lipid membranes. A slight structural difference in the sterol affects the membrane order significantly.

並列關鍵字

fluorescence microscopy ； sterol ； lipid ； membrane

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