Department of Biophysics, Medical College of Wisconsin, 8701 Watertown Plank Road, Milwaukee, WI, 53226, USA.
Exp Eye Res. 2021 May;206:108536. doi: 10.1016/j.exer.2021.108536. Epub 2021 Mar 12.
Eye lens membranes are complex biological samples. They consist of a variety of lipids that form the lipid bilayer matrix, integral proteins embedded into the lipid bilayer, and peripheral proteins. This molecular diversity in membrane composition induces formation of lipid domains with particular physical properties that are responsible for the maintenance of proper membrane functions. These domains can be, and have been, effectively described in terms of the rotational diffusion of lipid spin labels and oxygen collision with spin labels using the saturation recovery (SR) electron paramagnetic resonance method and, now, using stretched exponential function for the analysis of SR signals. Here, we report the application of the stretched exponential function analysis of SR electron paramagnetic resonance signals coming from cholesterol analog, androstane spin label (ASL) in the lipid bilayer portion of intact fiber cell plasma membranes (IMs) isolated from the cortex and nucleus of porcine eye lenses. Further, we compare the properties of these IMs with model lens lipid membranes (LLMs) derived from the total lipids extracted from cortical and nuclear IMs. With this approach, the IM can be characterized by the continuous probability density distribution of the spin-lattice relaxation rates associated with the rotational diffusion of a spin label, and by the distribution of the oxygen transport parameter within the IM (i.e., the collision rate of molecular oxygen with the spin label). We found that the cortical and nuclear LLMs possess very different, albeit homogenous, spin lattice relaxation rates due to the rotational diffusion of ASL, indicating that the local rigidity around the spin label in nuclear LLMs is considerably greater than that in cortical LLMs. However, the oxygen transport parameter around the spin label is very similar and slightly heterogenous for LLMs from both sources. This heterogeneity was previously missed when distinct exponential analysis was used. The spin lattice relaxation rates due to either the rotational diffusion of ASL or the oxygen collision with the spin label in nuclear IMs have slower values and wider distributions compared with those of cortical IMs. From this evidence, we conclude that lipids in nuclear IMs are less fluid and more heterogeneous than those in cortical membranes. Additionally, a comparison of properties of IMs with corresponding LLMs, and lipid and protein composition analysis, allow us to conclude that the decreased lipid-to-protein ratio not only induces greater rigidity of nuclear IMs, but also creates domains with the considerably decreased and variable oxygen accessibility. The advantages and disadvantages of this method, as well as its use for the cluster analysis, are discussed.
晶状体膜是复杂的生物样本。它们由多种形成脂质双层基质的脂质、嵌入脂质双层的整合蛋白和外周蛋白组成。这种膜组成的分子多样性诱导具有特定物理性质的脂质域的形成，这些物理性质负责维持适当的膜功能。这些域可以而且已经可以使用脂质旋转标记的氧碰撞和旋转标记的饱和恢复 (SR) 电子顺磁共振方法有效地描述，现在，使用拉伸指数函数来分析 SR 信号。在这里，我们报告了拉伸指数函数分析胆固醇类似物、雄烷自旋标记 (ASL) 在从猪眼晶状体皮质和核分离的完整纤维细胞质膜 (IM) 的脂质双层部分中的 SR 电子顺磁共振信号的应用。此外，我们比较了这些 IM 的性质与源自皮质和核 IM 总脂质的模型晶状体脂质膜 (LLM)。通过这种方法，IM 可以通过与旋转扩散相关的自旋晶格弛豫率的连续概率密度分布来表征，并且可以通过 IM 内的氧传输参数分布来表征（即分子氧与自旋标记的碰撞速率）。我们发现，皮质和核 LLM 具有非常不同的，尽管是均匀的，自旋晶格弛豫率由于 ASL 的旋转扩散，这表明核 LLM 中自旋标记周围的局部刚性比皮质 LLM 大得多。然而，来自两个来源的 LLM 中，自旋标记周围的氧传输参数非常相似且略有不均匀。当使用不同的指数分析时，这种异质性以前被忽略了。与皮质 IM 相比，核 IM 中由于 ASL 的旋转扩散或自旋标记与氧的碰撞引起的自旋晶格弛豫率具有较慢的值和较宽的分布。从这些证据中，我们得出结论，核 IM 中的脂质比皮质膜中的脂质流动性更小且异质性更大。此外，将 IM 的性质与相应的 LLM、脂质和蛋白质组成分析进行比较，使我们能够得出结论，脂质与蛋白质的比例降低不仅会增加核 IM 的刚性，还会产生具有显著降低和可变氧可及性的域。讨论了这种方法的优缺点以及用于聚类分析的方法。
