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. 2002 Jan;82(1 Pt 1):285–298. doi: 10.1016/S0006-3495(02)75394-0

Molecular organization of cholesterol in polyunsaturated membranes: microdomain formation.

Michael R Brzustowicz 1, Vadim Cherezov 1, Martin Caffrey 1, William Stillwell 1, Stephen R Wassall 1
PMCID: PMC1302469  PMID: 11751316

Abstract

The molecular organization of cholesterol in phospholipid bilayers composed of 1,2-diarachidonylphosphatidylcholine (20:4-20:4PC), 1-stearoyl-2-arachidonylphosphatidylcholine (18:0-20:4PC), and 20:4-20:4PC/18:0-20:4PC (1/1 mol) was investigated by solid-state (2)H NMR and by low- and wide-angle x-ray diffraction (XRD). On the basis of distinct quadrupolar powder patterns arising from [3 alpha-(2)H(1)]cholesterol intercalated into the membrane and phase separated as solid, solubility chi(NMR)(chol) = 17 +/- 2 mol% and tilt angle alpha(0) = 25 +/- 1 degrees in 20:4-20:4PC were determined. The corresponding values in 18:0-20:4PC were chi (NMR)(chol) > or = 50 mol% and alpha(0) = 16 +/- 1 degrees. Cholesterol solubility determined by XRD was chi(NMR)(chol) = 15 +/- 2 mol% and chi(NMR)(chol) = 49 +/- 1 mol% for 20:4-20:4PC and 18:0-20:4PC, respectively. XRD experiments show that the solid sterol is monohydrate crystals presumably residing outside the bilayer. The (2)H NMR spectrum for equimolar [3 alpha-(2)H(1)]cholesterol added to mixed 20:4-20:4PC/18:0-20:4PC (1/1 mol) membranes is consistent with segregation of cholesterol into 20:4-20:4PC and 18:0-20:4PC microdomains of <160 A in size that preserve the molecular organization of sterol in the individual phospholipid constituents. Our results demonstrate unambiguously that cholesterol has low affinity to polyunsaturated fatty acids and support hypotheses of lateral phase separation of membrane constituents into sterol-poor/polyunsaturated fatty acid-rich and sterol-rich/saturated fatty acid-rich microdomains.

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Selected References

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  1. Albert A. D., Boesze-Battaglia K., Paw Z., Watts A., Epand R. M. Effect of cholesterol on rhodopsin stability in disk membranes. Biochim Biophys Acta. 1996 Sep 13;1297(1):77–82. doi: 10.1016/0167-4838(96)00102-1. [DOI] [PubMed] [Google Scholar]
  2. Applegate K. R., Glomset J. A. Computer-based modeling of the conformation and packing properties of docosahexaenoic acid. J Lipid Res. 1986 Jun;27(6):658–680. [PubMed] [Google Scholar]
  3. Bloom M., Thewalt J. L. Time and distance scales of membrane domain organization. Mol Membr Biol. 1995 Jan-Mar;12(1):9–13. doi: 10.3109/09687689509038489. [DOI] [PubMed] [Google Scholar]
  4. Brown D. A., London E. Structure and function of sphingolipid- and cholesterol-rich membrane rafts. J Biol Chem. 2000 Jun 9;275(23):17221–17224. doi: 10.1074/jbc.R000005200. [DOI] [PubMed] [Google Scholar]
  5. Brown M. F. Modulation of rhodopsin function by properties of the membrane bilayer. Chem Phys Lipids. 1994 Sep 6;73(1-2):159–180. doi: 10.1016/0009-3084(94)90180-5. [DOI] [PubMed] [Google Scholar]
  6. Brzustowicz M. R., Stillwell W., Wassall S. R. Molecular organization of cholesterol in polyunsaturated phospholipid membranes: a solid state 2H NMR investigation. FEBS Lett. 1999 May 21;451(2):197–202. doi: 10.1016/s0014-5793(99)00567-0. [DOI] [PubMed] [Google Scholar]
  7. Craven B. M. Crystal structure of cholesterol monohydrate. Nature. 1976 Apr 22;260(5553):727–729. doi: 10.1038/260727a0. [DOI] [PubMed] [Google Scholar]
  8. Douliez J. P., Léonard A., Dufourc E. J. Restatement of order parameters in biomembranes: calculation of C-C bond order parameters from C-D quadrupolar splittings. Biophys J. 1995 May;68(5):1727–1739. doi: 10.1016/S0006-3495(95)80350-4. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Everts S., Davis J. H. 1H and (13)C NMR of multilamellar dispersions of polyunsaturated (22:6) phospholipids. Biophys J. 2000 Aug;79(2):885–897. doi: 10.1016/S0006-3495(00)76344-2. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Gliss C., Randel O., Casalta H., Sackmann E., Zorn R., Bayerl T. Anisotropic motion of cholesterol in oriented DPPC bilayers studied by quasielastic neutron scattering: the liquid-ordered phase. Biophys J. 1999 Jul;77(1):331–340. doi: 10.1016/S0006-3495(99)76893-1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Guo W., Hamilton J. A. A multinuclear solid-state NMR study of phospholipid-cholesterol interactions. Dipalmitoylphosphatidylcholine-cholesterol binary system. Biochemistry. 1995 Oct 31;34(43):14174–14184. doi: 10.1021/bi00043a023. [DOI] [PubMed] [Google Scholar]
  12. Hernandez-Borrell J., Keough K. M. Heteroacid phosphatidylcholines with different amounts of unsaturation respond differently to cholesterol. Biochim Biophys Acta. 1993 Dec 12;1153(2):277–282. doi: 10.1016/0005-2736(93)90416-w. [DOI] [PubMed] [Google Scholar]
  13. Huang J., Buboltz J. T., Feigenson G. W. Maximum solubility of cholesterol in phosphatidylcholine and phosphatidylethanolamine bilayers. Biochim Biophys Acta. 1999 Feb 4;1417(1):89–100. doi: 10.1016/s0005-2736(98)00260-0. [DOI] [PubMed] [Google Scholar]
  14. Huang J., Feigenson G. W. A microscopic interaction model of maximum solubility of cholesterol in lipid bilayers. Biophys J. 1999 Apr;76(4):2142–2157. doi: 10.1016/S0006-3495(99)77369-8. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Huang T. H., Lee C. W., Das Gupta S. K., Blume A., Griffin R. G. A 13C and 2H nuclear magnetic resonance study of phosphatidylcholine/cholesterol interactions: characterization of liquid-gel phases. Biochemistry. 1993 Dec 7;32(48):13277–13287. doi: 10.1021/bi00211a041. [DOI] [PubMed] [Google Scholar]
  16. Huster D., Arnold K., Gawrisch K. Influence of docosahexaenoic acid and cholesterol on lateral lipid organization in phospholipid mixtures. Biochemistry. 1998 Dec 8;37(49):17299–17308. doi: 10.1021/bi980078g. [DOI] [PubMed] [Google Scholar]
  17. Huster D., Jin A. J., Arnold K., Gawrisch K. Water permeability of polyunsaturated lipid membranes measured by 17O NMR. Biophys J. 1997 Aug;73(2):855–864. doi: 10.1016/S0006-3495(97)78118-9. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Jacob R. F., Cenedella R. J., Mason R. P. Direct evidence for immiscible cholesterol domains in human ocular lens fiber cell plasma membranes. J Biol Chem. 1999 Oct 29;274(44):31613–31618. doi: 10.1074/jbc.274.44.31613. [DOI] [PubMed] [Google Scholar]
  19. Kariel N., Davidson E., Keough K. M. Cholesterol does not remove the gel-liquid crystalline phase transition of phosphatidylcholines containing two polyenoic acyl chains. Biochim Biophys Acta. 1991 Feb 11;1062(1):70–76. doi: 10.1016/0005-2736(91)90336-7. [DOI] [PubMed] [Google Scholar]
  20. Loomis C. R., Shipley G. G., Small D. M. The phase behavior of hydrated cholesterol. J Lipid Res. 1979 May;20(4):525–535. [PubMed] [Google Scholar]
  21. Marsan M. P., Muller I., Ramos C., Rodriguez F., Dufourc E. J., Czaplicki J., Milon A. Cholesterol orientation and dynamics in dimyristoylphosphatidylcholine bilayers: a solid state deuterium NMR analysis. Biophys J. 1999 Jan;76(1 Pt 1):351–359. doi: 10.1016/S0006-3495(99)77202-4. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. McCabe M. A., Wassall S. R. Rapid deconvolution of NMR powder spectra by weighted fast Fourier transformation. Solid State Nucl Magn Reson. 1997 Dec;10(1-2):53–61. doi: 10.1016/s0926-2040(97)00024-6. [DOI] [PubMed] [Google Scholar]
  23. Mitchell D. C., Litman B. J. Effect of cholesterol on molecular order and dynamics in highly polyunsaturated phospholipid bilayers. Biophys J. 1998 Aug;75(2):896–908. doi: 10.1016/S0006-3495(98)77578-2. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Monck M. A., Bloom M., Lafleur M., Lewis R. N., McElhaney R. N., Cullis P. R. Evidence for two pools of cholesterol in the Acholeplasma laidlawii strain B membrane: a deuterium NMR and DSC study. Biochemistry. 1993 Mar 30;32(12):3081–3088. doi: 10.1021/bi00063a020. [DOI] [PubMed] [Google Scholar]
  25. Morrow M. R., Davis P. J., Jackman C. S., Keough K. M. Thermal history alters cholesterol effect on transition of 1-palmitoyl-2-linoleoyl phosphatidylcholine. Biophys J. 1996 Dec;71(6):3207–3214. doi: 10.1016/S0006-3495(96)79514-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Murari R., Murari M. P., Baumann W. J. Sterol orientations in phosphatidylcholine liposomes as determined by deuterium NMR. Biochemistry. 1986 Mar 11;25(5):1062–1067. doi: 10.1021/bi00353a017. [DOI] [PubMed] [Google Scholar]
  27. Oldfield E., Meadows M., Rice D., Jacobs R. Spectroscopic studies of specifically deuterium labeled membrane systems. Nuclear magnetic resonance investigation of the effects of cholesterol in model systems. Biochemistry. 1978 Jul 11;17(14):2727–2740. doi: 10.1021/bi00607a006. [DOI] [PubMed] [Google Scholar]
  28. Petersen N. O., Chan S. I. More on the motional state of lipid bilayer membranes: interpretation of order parameters obtained from nuclear magnetic resonance experiments. Biochemistry. 1977 Jun 14;16(12):2657–2667. doi: 10.1021/bi00631a012. [DOI] [PubMed] [Google Scholar]
  29. Phillips M. C. Cholesterol packing, crystallization and exchange properties in phosphatidylcholine vesicle systems. Hepatology. 1990 Sep;12(3 Pt 2):75S–82S. [PubMed] [Google Scholar]
  30. Polozova A., Litman B. J. Cholesterol dependent recruitment of di22:6-PC by a G protein-coupled receptor into lateral domains. Biophys J. 2000 Nov;79(5):2632–2643. doi: 10.1016/S0006-3495(00)76502-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Rajamoorthi K., Brown M. F. Bilayers of arachidonic acid containing phospholipids studied by 2H and 31P NMR spectroscopy. Biochemistry. 1991 Apr 30;30(17):4204–4212. doi: 10.1021/bi00231a015. [DOI] [PubMed] [Google Scholar]
  32. Rankin S. E., Addona G. H., Kloczewiak M. A., Bugge B., Miller K. W. The cholesterol dependence of activation and fast desensitization of the nicotinic acetylcholine receptor. Biophys J. 1997 Nov;73(5):2446–2455. doi: 10.1016/S0006-3495(97)78273-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Richter F., Rapp G., Finegold L. Miscibility gap in fluid dimyristoylphosphatidylcholine:cholesterol as "seen" by x rays. Phys Rev E Stat Nonlin Soft Matter Phys. 2001 Apr 26;63(5 Pt 1):051914–051914. doi: 10.1103/PhysRevE.63.051914. [DOI] [PubMed] [Google Scholar]
  34. Rietveld A., Simons K. The differential miscibility of lipids as the basis for the formation of functional membrane rafts. Biochim Biophys Acta. 1998 Nov 10;1376(3):467–479. doi: 10.1016/s0304-4157(98)00019-7. [DOI] [PubMed] [Google Scholar]
  35. Ruocco M. J., Siminovitch D. J., Long J. R., Das Gupta S. K., Griffin R. G. 2H and 13C nuclear magnetic resonance study of N-palmitoylgalactosylsphingosine (cerebroside)/cholesterol bilayers. Biophys J. 1996 Oct;71(4):1776–1788. doi: 10.1016/S0006-3495(96)79378-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Saiz L., Klein M. L. Structural properties of a highly polyunsaturated lipid bilayer from molecular dynamics simulations. Biophys J. 2001 Jul;81(1):204–216. doi: 10.1016/S0006-3495(01)75692-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Schindler H., Seelig J. Deuterium order parameters in relation to thermodynamic properties of a phospholiped bilayer. A statistical mechanical interpretation. Biochemistry. 1975 Jun 3;14(11):2283–2287. doi: 10.1021/bi00682a001. [DOI] [PubMed] [Google Scholar]
  38. Seelig J. 31P nuclear magnetic resonance and the head group structure of phospholipids in membranes. Biochim Biophys Acta. 1978 Jul 31;515(2):105–140. doi: 10.1016/0304-4157(78)90001-1. [DOI] [PubMed] [Google Scholar]
  39. Shieh H. S., Hoard L. G., Nordman C. E. Crystal structure of anhydrous cholesterol. Nature. 1977 May 19;267(5608):287–289. doi: 10.1038/267287a0. [DOI] [PubMed] [Google Scholar]
  40. Simons K., Ikonen E. Functional rafts in cell membranes. Nature. 1997 Jun 5;387(6633):569–572. doi: 10.1038/42408. [DOI] [PubMed] [Google Scholar]
  41. Smaby J. M., Momsen M. M., Brockman H. L., Brown R. E. Phosphatidylcholine acyl unsaturation modulates the decrease in interfacial elasticity induced by cholesterol. Biophys J. 1997 Sep;73(3):1492–1505. doi: 10.1016/S0006-3495(97)78181-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Sömjen G. J., Lipka G., Schulthess G., Koch M. H., Wachtel E., Gilat T., Hauser H. Behavior of cholesterol and spin-labeled cholestane in model bile systems studied by electron spin resonance and synchrotron x-ray. Biophys J. 1995 Jun;68(6):2342–2349. doi: 10.1016/S0006-3495(95)80416-9. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Taylor M. G., Akiyama T., Saitô H., Smith I. C. Direct observation of the properties of cholesterol in membranes by deuterium NMR. Chem Phys Lipids. 1982 Dec;31(4):359–379. doi: 10.1016/0009-3084(82)90072-x. [DOI] [PubMed] [Google Scholar]
  44. Thewalt J. L., Bloom M. Phosphatidylcholine: cholesterol phase diagrams. Biophys J. 1992 Oct;63(4):1176–1181. doi: 10.1016/S0006-3495(92)81681-8. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Vist M. R., Davis J. H. Phase equilibria of cholesterol/dipalmitoylphosphatidylcholine mixtures: 2H nuclear magnetic resonance and differential scanning calorimetry. Biochemistry. 1990 Jan 16;29(2):451–464. doi: 10.1021/bi00454a021. [DOI] [PubMed] [Google Scholar]
  46. Zhu T., Caffrey M. Thermodynamic, thermomechanical, and structural properties of a hydrated asymmetric phosphatidylcholine. Biophys J. 1993 Aug;65(2):939–954. doi: 10.1016/S0006-3495(93)81108-1. [DOI] [PMC free article] [PubMed] [Google Scholar]

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