Cannon Brian, Lewis Anthony, Metze Jennifer, Thiagarajan Visveswaran, Vaughn Mark W, Somerharju Pentti, Virtanen Jorma, Huang Juyang, Cheng Kwan Hon
Department of Physics, Texas Tech University, Lubbock, Texas 79409-1051, USA.
J Phys Chem B. 2006 Mar 30;110(12):6339-50. doi: 10.1021/jp0558371.
Fluorescence and Fourier transform infrared (FTIR) spectroscopic techniques were used to explore the effect of added cholesterol on the composition-dependent formation of putative phospholipid headgroup superlattices in fluid 1-palmitoyl-2-oleoyl-phosphatidylethanolamine/1-palmitoyl-2-oleoyl-phosphatidylcholine/cholesterol (POPE/POPC/CHOL) bilayers. Steady-state fluorescence anisotropy measurements of diphenylhexatriene (DPH) chain-labeled phosphatidylcholine (DPH-PC) revealed significant dips at several POPE-to-phospholipid mole fractions (X(PE)'s) when the cholesterol-to-lipid mole fraction (X(CHOL)) was fixed at 0.00, 0.35, 0.40, and 0.50. Most of the observed dips occur at or close to critical X(PE)'s predicted by the Headgroup Superlattice (SL) model, suggesting that phospholipid headgroups of different structures tend to adopt regular distributions even in the presence of cholesterol. Time-resolved fluorescence anisotropy measurements revealed that DPH-PC senses a disordered and highly mobile microenvironment in the POPE/POPC/CHOL bilayers at those critical X(PE)'s, indicating that this probe may partition to defect regions in the bilayers. The presence of coexisting packing defect regions and regularly distributed SL domains is a key feature predicted by the Headgroup SL model. Importantly, probe-free FTIR measurements of acyl chain C-H, interfacial carbonyl, and headgroup phosphate stretching peak frequencies revealed the presence of abrupt changes at X(PE)'s close to those observed in the fluorescence data. When X(PE) was varied from 0.60 to 0.72 and X(CHOL) from 0.34 to 0.46, a clear dip at the lipid composition coordinates (X(PE), X(CHOL)) approximately (0.68, 0.40) was observed in the three-dimensional surface plots of DPH-PC anisotropy as well as the carbonyl and phosphate stretching frequencies. The critical X(CHOL) at 0.40 agrees with the Cholesterol SL model, which assumes that cholesterol and phospholipid form SL domains at the lipid acyl chain level. In conclusion, this study provides evidence that cholesterol supports formation of phospholipid headgroup SLs in fluid state ternary lipid bilayers. The feasibility of the parallel existence of SLs at the lipid headgroup and acyl chain levels supports the relevance of the lipid SL model for the membranes of eukaryotic cells that typically contain significant amounts of cholesterol. We speculate that lipid SL formation may play a central role in the regulation of membrane lipid compositions, maintenance of organelle boundaries, and other crucial phenomena in those cells.
利用荧光和傅里叶变换红外（FTIR）光谱技术，研究了添加胆固醇对流体态1-棕榈酰-2-油酰磷脂酰乙醇胺/1-棕榈酰-2-油酰磷脂酰胆碱/胆固醇（POPE/POPC/CHOL）双层膜中假定的磷脂头基超晶格组成依赖性形成的影响。对二苯基己三烯（DPH）链标记的磷脂酰胆碱（DPH-PC）进行稳态荧光各向异性测量发现，当胆固醇与脂质的摩尔分数（X(CHOL)）固定为0.00、0.35、0.40和0.50时，在几个POPE与磷脂的摩尔分数（X(PE)）处出现了显著的下降。观察到的大多数下降发生在头基超晶格（SL）模型预测的临界X(PE)处或其附近，这表明即使在存在胆固醇的情况下，不同结构的磷脂头基也倾向于采用规则分布。时间分辨荧光各向异性测量表明，在这些临界X(PE)处，DPH-PC在POPE/POPC/CHOL双层膜中感受到无序且高度可移动的微环境，这表明该探针可能分配到双层膜中的缺陷区域。共存的堆积缺陷区域和规则分布的SL域的存在是头基SL模型预测的一个关键特征。重要的是，对酰基链C-H、界面羰基和头基磷酸酯伸缩峰频率进行的无探针FTIR测量表明，在接近荧光数据中观察到的X(PE)处存在突然变化。当X(PE)从0.60变化到0.72且X(CHOL)从0.34变化到0.46时，在DPH-PC各向异性以及羰基和磷酸酯伸缩频率的三维表面图中，在脂质组成坐标（X(PE)，X(CHOL)）约为（0.68，0.40）处观察到明显的下降。0.40处的临界X(CHOL)与胆固醇SL模型一致，该模型假设胆固醇和磷脂在脂质酰基链水平形成SL域。总之，本研究提供了证据表明胆固醇支持流体态三元脂质双层膜中磷脂头基SL的形成。脂质头基和酰基链水平上SL平行存在的可行性支持了脂质SL模型与典型含有大量胆固醇的真核细胞膜的相关性。我们推测脂质SL的形成可能在调节膜脂质组成、维持细胞器边界以及这些细胞中的其他关键现象中发挥核心作用。
