Haque M E, McIntosh T J, Lentz B R
Department of Biochemistry & Program in Molecular/Cell Biophysics, University of North Carolina, Chapel Hill, North Carolina 27599-7260, USA.
Biochemistry. 2001 Apr 10;40(14):4340-8. doi: 10.1021/bi002030k.
Poly(ethylene glycol) (PEG)-mediated fusion of phosphatidylcholine model membranes has been shown to mimic the protein-mediated biomembrane process [Lee, J., and Lentz, B. R. (1998) Proc. Natl. Acad. Sci. U.S.A. 95, 9274-9279]. Unlike the simple model membranes used in this earlier study, the lipid composition of fusogenic biomembranes is quite complex. The purpose of this paper was to examine PEG-mediated fusion of highly curved (SUV) and largely uncurved (LUV) membrane vesicles composed of different lipids in order to identify lipid compositions that produce highly fusogenic membranes. Starting with liposomes composed of five lipids with different physical properties, dioleoylphosphatidylcholine (DOPC), dioleoylphosphatidylethanolamine (DOPE), dioleoylphosphatidylserine (DOPS), bovine brain sphingomyelin (SM), and cholesterol (CH), we systematically varied the composition and tested for the extent of PEG-mediated fusion after 5 min of treatment. We found that a vesicle system composed of four lipids, DOPC/DOPE/SM/CH, fused optimally at a 35/30/15/20 molar ratio. Each lipid seemed to play a part in optimizing the membrane for fusion. PE disrupted outer leaflet packing as demonstrated with TMA-DPH lifetime, C(6)-NBD-PC partitioning, and DPH anisotropy measurements, and thus significantly enhanced fusion and rupture, without significantly altering interbilayer approach (X-ray diffraction). An optimal ratio of PC/PE (35/30) produced a balance between fusion and rupture. CH and SM, when present at an optimal ratio of 3/4 in vesicles containing the optimal PC/PE ratio, reduced rupture without significantly reducing fusion. This optimal CH/SM ratio also enhanced outer leaflet packing, suggesting that fusion is dependent not only on outer leaflet packing but also on the properties of the inner leaflet. Addition of CH without SM enhanced rupture relative to fusion, while SM alone reduced both rupture and fusion. The optimal lipid composition is very close to the natural synaptic vesicle composition, suggesting that the synaptic vesicle composition is optimized with respect to fusogenicity.
聚乙二醇（PEG）介导的磷脂酰胆碱模型膜融合已被证明可模拟蛋白质介导的生物膜过程[Lee, J., and Lentz, B. R. (1998) Proc. Natl. Acad. Sci. U.S.A. 95, 9274 - 9279]。与该早期研究中使用的简单模型膜不同，促融合生物膜的脂质组成相当复杂。本文的目的是研究PEG介导的由不同脂质组成的高度弯曲（小单层囊泡，SUV）和基本不弯曲（大单层囊泡，LUV）膜囊泡的融合，以确定产生高度促融合膜的脂质组成。我们从由具有不同物理性质的五种脂质组成的脂质体开始，即二油酰磷脂酰胆碱（DOPC）、二油酰磷脂酰乙醇胺（DOPE）、二油酰磷脂酰丝氨酸（DOPS）、牛脑鞘磷脂（SM）和胆固醇（CH），系统地改变其组成，并在处理5分钟后测试PEG介导的融合程度。我们发现由四种脂质DOPC/DOPE/SM/CH组成的囊泡系统在摩尔比为35/30/15/20时融合最佳。每种脂质似乎都在优化膜融合方面发挥作用。PE破坏了外叶层堆积，这通过TMA - DPH寿命、C(6) - NBD - PC分配和DPH各向异性测量得到证明，因此显著增强了融合和破裂，而没有显著改变双层间距离（X射线衍射）。PC/PE的最佳比例（35/30）在融合和破裂之间产生了平衡。当CH和SM以最佳比例3/4存在于含有最佳PC/PE比例的囊泡中时，减少了破裂而没有显著降低融合。这种最佳的CH/SM比例也增强了外叶层堆积，表明融合不仅取决于外叶层堆积，还取决于内叶层的性质。相对于融合，添加CH而不添加SM会增强破裂，而单独的SM会降低破裂和融合。最佳脂质组成非常接近天然突触小泡的组成，这表明突触小泡的组成在促融合性方面是经过优化的。
