Ralph N. Adams Institute for Bioanalytical Chemistry, University of Kansas, 2030 Becker Drive, Lawrence, Kansas 66047, USA.
J Phys Chem B. 2013 Jul 3;117(26):7959-66. doi: 10.1021/jp405312a. Epub 2013 Jun 24.
Single molecule fluorescence measurements are used to probe the effects of GM1 in DPPC monolayers. Langmuir-Blodgett films of GM1 and DPPC were doped with ~10(-8) mol % of the fluorescent lipid probe, BODIPY-PC, and transferred onto glass substrates at 23 mN/m. As shown previously, the individual orientation of each BODIPY-PC probe in the membrane can be measured using defocused polarized total internal reflection fluorescence microscopy, revealing changes in film properties at the molecular level. Here, BODIPY-PC tilt angle histograms are used to characterize the effects of GM1 in DPPC films from 0.05 to 100 mol % GM1. At high GM1 levels (>5 mol % GM1), trends in the single molecule measurements agree with previous bulk measurements showing the turnover from condensing to expanding influence of GM1 at 15-20 mol %, thus validating the single molecule approach. At biologically relevant, low concentrations of GM1 (<5 mol % GM1), where bulk fluorescence measurements are less informative, the single molecule measurements reveal a marked influence of GM1 on film properties. The addition of trace amounts of GM1 to DPPC films leads to an expansion of the film which continues to 0.10 mol % GM1, above which the trend reverses and the condensing effect previously noted is observed.
单分子荧光测量用于探测 GM1 在 DPPC 单层中的作用。GM1 和 DPPC 的 Langmuir-Blodgett 薄膜中掺杂了约 10(-8) mol % 的荧光脂质探针 BODIPY-PC，并在 23 mN/m 的压力下转移到玻璃基底上。如前所述，使用离焦偏振全内反射荧光显微镜可以测量每个 BODIPY-PC 探针在膜中的单个取向，从而揭示分子水平上的膜性质变化。在这里，BODIPY-PC 倾斜角直方图用于表征 GM1 在 DPPC 薄膜中的作用，GM1 的浓度范围为 0.05 至 100 mol %。在 GM1 高浓度 (>5 mol % GM1)下，单分子测量的趋势与先前的体相测量一致，表明 GM1 在 15-20 mol %时从凝结到扩展的转变，从而验证了单分子方法的有效性。在生物学相关的低 GM1 浓度 (<5 mol % GM1)下，体相荧光测量的信息量较少，单分子测量揭示了 GM1 对膜性质的显著影响。向 DPPC 薄膜中添加痕量 GM1 会导致膜扩张，直到 GM1 的浓度达到 0.10 mol %，之后趋势反转，之前观察到的凝结效应再次出现。
