Effect of pulmonary surfactant protein SP-B on the micro- and nanostructure of phospholipid films
Effect of pulmonary surfactant protein SP-B on the micro- and nanostructure of phospholipid films
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DOI：
10.1016/s0006-3495(04)74106-5
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发表时间：
2004-01-01
影响因子：
3.4
通讯作者：
Pérez-Gil, J
中科院分区：
文献类型：
--
作者：
Cruz, A;Vázquez, L;Pérez-Gil, J
将二棕榈酰磷脂酰胆碱（DPPC）和DPPC/二棕榈酰磷脂酰甘油（DPPG）（7：3，w/w）在不存在或存在2、5、10或20重量%的猪表面活性剂蛋白SP-B的情况下的单层在表面天平的空气-液体界面处展开，在等温线的液体膨胀/液体冷凝（LE-LC）平台中压缩至表面压力，转移到云母载体上，并通过扫描力显微镜进行分析。在没有蛋白质的情况下，薄膜显示微米级的凝聚域的形态和大小，类似于原位观察到的那些在空气-液体界面的落射荧光显微镜。扫描力显微镜允许检查的共存相在一个更高的分辨率比以前实现的荧光显微镜。DPPC膜的LE区和LC区本质上是异质的。LC微域包含许多扩展样岛，而显然是液体扩展的区域被相互连接的纳米域的缩合样框架覆盖。肺表面活性物质蛋白SP-B的存在影响的LE和LC区域的DPPC和DPPC/DPPG膜的微观和纳米水平的分布。凝聚的微区变得越来越多，但它们的尺寸减小，导致DPPC和DPPC/DPPG膜中的总LC相的量的总体减少。在纳米级水平上，SP-B还引起LE相中的凝聚状纳米畴的尺寸的显著减小和LE/LC界面的长度的增加。SP-B促进a.脂质和脂质-蛋白质纳米结构域的纳米级框架，其与在膜转移和操作期间观察到的对膜变形和破裂的实质性机械阻力相关。SP-B对脂质膜的纳米级结构的影响在DPPC/DPPG中比在纯DPPC膜中更大，表明静电脂质-蛋白质相互作用的额外贡献。由SP-B的磷脂膜的纳米级结构的改变提供了蛋白质的结构框架，同时维持结构稳定性以及在由呼吸力学施加的极端条件下的表面活性剂膜的动态灵活性。SP-B也形成了分离的二维簇，与DPPC和DPPC/DPPG膜的LC微区和LE区域之间的边界相关联。这些簇以高于2%重量的蛋白质-脂质比例存在表明表面活性剂脂质-蛋白质复合物中SP-B的浓度可能接近脂质膜中蛋白质的溶解度极限。
Monolayers of dipalmitoylphosphatidylcholine (DPPC) and DPPC/dipalmitoylphosphatidylglycerol (DPPG) (7:3, w/w) in the absence or in the presence of 2, 5, 10, or 20 weight percent of porcine surfactant protein SP-B were spread at the air-liquid interface of a surface balance, compressed up to surface pressures in the liquid-expanded/liquid-condensed (LE-LC) plateau of the isotherm, transferred onto mica supports, and analyzed by scanning force microscopy. In the absence of protein, the films showed micrometer-sized condensed domains with morphology and size that were analogous to those observed in situ at the air-liquid interface by epifluorescence microscopy. Scanning force microscopy permits examination of the coexisting phases at a higher resolution than previously achieved with fluorescent microscopy. Both LE and LC regions of DPPC films were heterogeneous in nature. LC microdomains contained numerous expanded-like islands whereas regions apparently liquid-expanded were covered by a condensed-like framework of interconnected nanodomains. Presence of increasing amounts of pulmonary surfactant protein SP-B affected the distribution of the LE and LC regions of DPPC and DPPC/DPPG films both at the microscopic and the nanoscopic level. The condensed microdomains became more numerous but their size decreased, resulting in an overall reduction of the amount of total LC phase in both DPPC and DPPC/DPPG films. At the nanoscopic level, SP-B also caused a marked reduction of the size of the condensed-like nanodomains in the LE phase and an increase in the length of the LE/LC interface. SP-B promotes a. ne nanoscopic framework of lipid and lipid-protein nanodomains that is associated with a substantial mechanical resistance to film deformation and rupture as observed during film transference and manipulation. The effect of SP-B on the nanoscopic structure of the lipid films was greater in DPPC/DPPG than in pure DPPC films, indicating additional contributions of electrostatic lipid-protein interactions. The alterations of the nanoscopic structures of phospholipid films by SP-B provide the structural framework for the protein simultaneously sustaining structural stability as well as dynamical flexibility in surfactant films at the extreme conditions imposed by the respiratory mechanics. SP-B also formed segregated two-dimensional clusters that were associated with the boundaries between LC microdomains and the LE regions of DPPC and DPPC/DPPG films. The presence of these clusters at protein-to-lipid proportions above 2% by weight suggests that the concentration of SP-B in the surfactant lipid-protein complexes may be close to the solubility limit of the protein in the lipid films.