High-pressure freezing causes structural alterations in phospholipid model membranes
High-pressure freezing causes structural alterations in phospholipid model membranes
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DOI：
10.1046/j.1365-2818.1998.00328.x
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发表时间：
1998-06-01
影响因子：
2
通讯作者：
Meyer, HW
中科院分区：
文献类型：
--
作者：
Semmler, K;Wunderlich, J;Meyer, HW
研究了高压冷冻（HPF）对磷脂模型膜中脂质排列的影响。用冷冻断裂电镜分析了由纯双棕榈酰磷脂酰胆碱（DPPC）和DPPC与支链磷脂胆碱（1,2 -二（4-十二烷基棕榈酰）- n-甘油-3-磷脂胆碱）混合而成的脂质体。用液态丙烷或HPE冷冻脂质体，在高压条件下冷冻时，双组分脂质体体系的特征大波纹相的形态发生了巨大变化。通过对照实验排除了乙醇作为压力传递介质的影响。相比之下，没有观察到纯DPPC双层膜的高压改变。我们假设二元体系的修饰是由于压力引起的应力和不稳定的脂质分子包装结构的松弛。采用新设计的夹在铜片上的样品架和高压冷冻机Balzers HPMO10进行高压冷冻。对于流体样品的冻裂，夹层结构优于原来的支架系统。通过在支架之间插入垫片，可以对厚度为20-100 μ m的样品进行高压冷冻。夹心支架配有热电偶，以监测冷却速率，并允许精确的样品温度控制。尽管与原来的支架相比，质量减少了两倍，但HPF冷却速度没有提高（4000度Cs-1）。结果表明，高压冷冻过程中的冷却过程主要由冷媒速度决定。
The influence of high-pressure freezing (HPF) on the lipid arrangement in phospholipid model membranes has been investigated. Liposomes consisting of pure dipalmitoylphosphatidylcholine (DPPC) and of DPPC mixed with a branched-chain phosphocholine ( 1,2 -di(4-dodecyl-palmitoyl)-sn-glycero-3-phosphocholine) have been analysed by freeze-fracture electron microscopy. The liposomes were frozen either by plunging into liquid propane or by HPE The characteristic macroripple-phase of the two-component liposome system is drastically changed in its morphology when frozen under high-pressure conditions. The influence of ethanol which acts as pressure transfer medium was ruled out by control experiments. In contrast, no high-pressure alterations of the pure DPPC bilayer membrane have been observed. We assume that the modification of the binary system is due to a pressure-induced relaxation of a stressed and unstable lipid molecule packing configuration. HPF was performed with a newly designed sample holder for using sandwiched copper platelets with the high-pressure freezing machine Balzers HPMO10. The sandwich construction turned out to be superior to the original holder system with regard to freeze-fracturing of fluid samples. By inserting a spacer between the supports samples with a thickness of 20-100 mu m can be high-pressure frozen. The sandwich holder is provided with a thermocouple to monitor cooling rates and allows exact sample temperature control. Despite a two-fold mass reduction compared to the original holder no HPF cooling rate improvement has been achieved (4000 degrees Cs-1). We conclude that the cooling process in high-pressure freezing is determined mainly by cryogen velocity.