High-pressure freezing causes structural alterations in phospholipid model membranes.
The influence of high-pressure freezing (HPF) on the lipid arrangement in phospholipid model membranes has been investigated. Liposomes consisting of pure dipalmitoyl-phosphatidylcholine (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 HPF. 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 HPM010. 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 microns 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.
Semmler
K
K
Institute of Ultrastructure Research, Friedrich-Schiller-University Jena, Germany.
Wunderlich
J
J
Richter
W
W
Meyer
H W
HW
D015060:1,2-Dipalmitoylphosphatidylcholine / Q000032:analysis; D015925:Cryopreservation / Q000379:methods*; D005614:Freeze Fracturing / Q000379:methods*; D008051:Lipid Bilayers; D008081:Liposomes; D008854:Microscopy, Electron; D010743:Phospholipids / Q000737:chemistry*; D011312:Pressure; D014961:X-Ray Diffraction
D008051:Lipid Bilayers; D008081:Liposomes; D010743:Phospholipids; D015060:1,2-Dipalmitoylphosphatidylcholine