Electron spin resonance evidence for vertical asymmetry in animal cell membranes.
Electron spin resonance evidence for vertical asymmetry in animal cell membranes.
复制标题
动物细胞膜垂直不对称性的电子自旋共振证据。
作者：
B. Wisnieski;K. Iwata
两个电子自旋共振（ESR）自旋标记用于监测细菌和动物细胞膜的物理状态：5 N10，癸烷的氮氧化物衍生物，和12 NS-GA，12-氮氧化物硬脂酸的葡糖胺衍生物。从大约5 ℃到45 ℃，以1 ℃的间隔记录光谱。通过分别测量烃和水信号的振幅，hH和hP，获得log hH/hP对1/K的Arrhenius图。两个不连续的Arrhenius图（在特征温度t1和th）观察到与细菌细胞膜的自旋标签独立。密封的动物细胞膜样品的分析揭示了当使用疏水性自旋标记物5 N10时的四个特征温度，但当使用两亲性自旋标记物12 NS-GA时仅两个。用12 NS-GA揭示的特征温度的具体设置取决于膜制备是由内而外（ISO）还是正面朝外（RSO）。纽卡斯尔病病毒是来自宿主的RSO质膜的来源，对该病毒的分析揭示了两个特征温度，约为14和33摄氏度。吞噬体是来源于LM细胞的ISO质膜的来源，对吞噬体的分析揭示了大约23和38 ℃的两个特征温度。当未密封或破坏的膜制备物用12 NS-GA自旋标记时，揭示了两组（RSO和ISO）特征温度。结果表明，动物细胞膜的内部和外部单层在物理上是不同的，并且糖基化的自旋标记物12 NS-GA在其翻转穿过膜双层的能力方面明显受到限制。在这项研究中，通过计算机分析ESR光谱数据来确定特征温度。
Two electron spin resonance (ESR) spin labels were used to monitor the physical state of bacterial and animal cell membranes: 5N10, a nitroxide derivative of decane, and 12NS-GA, a glucosamine derivative of 12-nitroxide stearic acid. Spectra were recorded at 1 degrees C intervals from approximately 5 to 45 degrees C. Arrhenius plots of log hH/hP vs. 1/K were obtained by measuring the amplitudes of the hydrocarbon and water signals, hH and hP, respectively. Two discontinuities in the Arrhenius plot (at characteristic temperatures t1 and th) were observed with bacterial cell membranes independent of the spin label employed. Analysis of sealed animal cell membrane samples revealed four characteristic temperatures when the hydrophobic spin lable 5N10 was used, but only two when the amphiphilic spin label 12NS-GA was used. The specific set of characteristic temperatures revealed with 12NS-GA depended on whether the membrane preparation was inside out (ISO) or right side out (RSO). Analysis of Newcastle disease virus, a source of RSO plasma membrane derived from host, revealed two characteristic temperatures at approximately 14 and 33 degrees C. Analysis of phagosomes, a source of ISO plasma membrane derived from LM cells, revealed two characteristic temperatures at approximately 23 and 38 degrees C. When unsealed or disrupted membrane preparations were spin labeled with 12NS-GA, both sets (RSO and ISO) of characteristic temperatures were revealed. The results indicate that the inner and outer monolayers of animal cell membranes are physically distinct and that the glycosylated spin label, 12NS-GA, is apparently restricted in its ability to flip across the membrane bilayer. In this study, characteristic temperatures were pinpointed by computer analysis of the ESR spectral data.