Nag K, Taneva S G, Perez-Gil J, Cruz A, Keough K M
Department of Biochemistry, Memorial University of Newfoundland, St. John's, Canada.
Biophys J. 1997 Jun;72(6):2638-50. doi: 10.1016/S0006-3495(97)78907-0.
Hydrophobic pulmonary surfactant (PS) proteins B (SP-B) and C (SP-C) modulate the surface properties of PS lipids. Epifluorescence microscopy was performed on solvent-spread monolayers of fluorescently labeled porcine SP-B (R-SP-B, labeled with Texas Red) and SP-C (F-SP-C, labeled with fluorescein) in dipalmitoylphosphatidylcholine (DPPC) (at protein concentrations of 10 and 20 wt%, and 10 wt% of both) under conditions of cyclic compression and expansion. Matrix-assisted laser desorption/ionization (MALDI) spectroscopy of R-SP-B and F-SP-C indicated that the proteins were intact and labeled with the appropriate fluorescent probe. The monolayers were compressed and expanded for four cycles at an initial rate of 0.64 A2 x mol(-1) x s(-1) (333 mm2 x s x [-1]) up to a surface pressure pi approximately 65 mN/m, and pi-area per residue (pi-A) isotherms at 22 +/- 1 degrees C were obtained. The monolayers were microscopically observed for the fluorescence emission of the individual proteins present in the film lipid matrix, and their visual features were video recorded for image analysis. The pi-A isotherms of the DPPC/protein monolayers showed characteristic "squeeze out" effects at pi approximately 43 mN/m for R-SP-B and 55 mN/m for F-SP-C, as had previously been observed for monolayers of the native proteins in DPPC. Both proteins associated with the expanded (fluid) phase of DPPC monolayers remained in or associated with the monolayers at high pi (approximately 65 mN/m) and redispersed in the monolayer upon its reexpansion. At comparable pi and area/molecule of the lipid, the proteins reduced the amounts of condensed (gel-like) phase of DPPC monolayers, with F-SP-C having a greater effect on a weight basis than did R-SP-B. In any one of the lipid/protein monolayers the amounts of the DPPC in condensed phase were the same at equivalent pi during compression and expansion and from cycle to cycle. This indicated that only minor loss of components from these systems occurred between compression-expansion cycles. This study indicates that hydrophobic PS proteins associate with the fluid phase of DPPC in films, some proteins remain at high surface pressures in the films, and such lipid-protein films can still attain high pi during compression.
疏水性肺表面活性物质（PS）蛋白B（SP - B）和蛋白C（SP - C）可调节PS脂质的表面特性。在二棕榈酰磷脂酰胆碱（DPPC）中（蛋白浓度分别为10%和20%重量，以及两者均为10%重量），于循环压缩和膨胀条件下，对用荧光标记的猪SP - B（R - SP - B，用德克萨斯红标记）和SP - C（F - SP - C，用荧光素标记）的溶剂铺展单层进行了落射荧光显微镜观察。对R - SP - B和F - SP - C进行基质辅助激光解吸/电离（MALDI）光谱分析表明，这些蛋白保持完整且用适当的荧光探针进行了标记。单层以0.64 Ų×mol⁻¹×s⁻¹（333 mm²×s⁻¹）的初始速率压缩和膨胀四个循环，直至表面压力π约为65 mN/m，并在22 ± 1℃下获得每个残基的π - 面积（π - A）等温线。通过显微镜观察膜脂质基质中单个蛋白的荧光发射，并对其视觉特征进行视频记录以进行图像分析。DPPC/蛋白单层的π - A等温线在π约为43 mN/m（对于R - SP - B）和55 mN/m（对于F - SP - C）时显示出特征性的“挤出”效应，这与之前在DPPC中天然蛋白单层所观察到的情况相同。与DPPC单层的膨胀（流体）相相关的两种蛋白在高π（约65 mN/m）时仍保留在单层中或与单层相关联，并在单层重新膨胀时重新分散。在脂质的可比π和分子面积下，这些蛋白减少了DPPC单层中凝聚（凝胶状）相的量，以重量计，F - SP - C的影响比R - SP - B更大。在任何一种脂质/蛋白单层中，在压缩、膨胀过程以及从一个循环到下一个循环的等效π下，凝聚相中的DPPC量是相同的。这表明在这些系统的压缩 - 膨胀循环之间仅发生了少量成分损失。本研究表明，疏水性PS蛋白与膜中DPPC的流体相相关联，一些蛋白在膜中保持高表面压力，并且这种脂质 - 蛋白膜在压缩过程中仍可达到高π。
