The Journal of Membrane Biology

, Volume 85, Issue 1, pp 79–86 | Cite as

Thermotropic behavior of retinal rod membranes and dispersions of extracted phospholipids

  • George P. Miljanich
  • Michael F. Brown
  • Susan Mabrey-Gaud
  • Edward A. Dratz
  • Julian M. Sturtevant


High sensitivity, differential scanning calorimetry studies of vovine retinal rod outer segment (ROS) disk membranes and aqueous dispersions of the extracted ROS phospholipids have been performed. ROS disk membranes were found to exhibit a broad peak of excess heat capacity with a maximum at less than about 3°C, ascribable to a gel-to-liquid crystalline phase transition of traction of the phospholipids. A similar thermotropic transition was observed for aqueous dispersions of the total extracted and purified ROS phospholipids. Comparison of the results obtained for the dispersion of total ROS phospholipids to those of the purified head group fractions. suggests that the thermotropic behavior reffects a gel-to-liquid crystalline transition, leading to lateral phase separation, involving those phosphatidylcholine (PC) molecules containing saturated fatty acylchains, possibley together with the highest melting ROS phosphatidylethanolamine (PE) and phosphatidylserine (PS) components. The interpretation of the thermal behavior of the ROS disk membranes depends on whether the transition is assumed to derive from the ROS PC and/or PE/PS fractions, and whether the transbilayer arrangement of the ROS phospholipids is assumed to be symmetric or asymmetric. The calorimetric data can be simply explained in terms of an asymmetric distribution of the major ROS disk membrane phospholipids (G.P. Miljanich et al.,J. Membrane Biol.60:249–255, 1981). In this case, the transition would arise from the PE/PS fractions in the outer ROS disk membrane monolyer, and the anticipated transition from the PC in the inner monolayer would be broadened due to interaction with cholesterol. For the ROS membranes at higher temperatures, two additional, irreversible transitions are observed at 57 and 72°C, corresponding to the thermal denauturation of opsin and rhodopsin, respectively.

Key Words

differential scanning calorimetrý retinal rod membranes lipid phase transitions cholesterol membrane asymmetry proitein denaturation rhodopsin vision 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Albert, A.D., Yeagle, P.L. 1983. Phospholipid domains in bovine retinal ro outer segment disk membranes.Proc. Natl. Acad. Sci. USA 80:7188–7191PubMedGoogle Scholar
  2. Albon, N., Stutevant, J.M. 1978. Nature of the gel to liquid crystal transition of synthetic phosphatidylcholines.Proc. Natl. Acad. Sci. USA 75:2258–2260PubMedGoogle Scholar
  3. Applebury, M.L., Zuckerman, D.M., Lamola, A.A., Jovin, T.M. 1974. Rhodopsin. Purification and recombination with phospholipids assayed by the metarhodopsin I-metarhodopsin II transition.Biochemistry 13:3448–3458PubMedGoogle Scholar
  4. Blumej, A. 1980. Thermotropic behavior of phosphatidylethanolamine-cholesterol and phosphatidylethanolamine-phosphatidylcholine-cholesterol mixtures.Biochemistry 21:4908–4913Google Scholar
  5. Brown, M.F., Deese, A.J., Dratz, E.A. 1982. Proton, carbon-13, and phosphorus-31 NMR methods for the investigation of rhodopsin-lipid interactions in retinal rod outer segment membranes.Methods Enzymol. 81:709–728PubMedGoogle Scholar
  6. Brown, M.F., Miljanich, G.P., Dratz, E.A. 1977a. Proton spinlattice relaxation of retinal rod outer segment membranes and liposomes of extracted phospholipids.Proc. Natl. Acad. Sci. USA 74:1978–1982PubMedGoogle Scholar
  7. Brown, M.F., Miljanich, G.P., Dratz, E.A. 1977b. Interpretation of 100- and 360-MHz proton magnetic resonance spectra of retinal rod outer segment disk membranes.Biochemistry 16:2640–2648PubMedGoogle Scholar
  8. Brown, M.F., Miljanich, G.P., Franklin, L.K., Dratz, E.A. 1976. H-NMR studies of protein-lipid interactions in retinal rod outer segment disc membranes.FEBS Lett. 70:56–60PubMedGoogle Scholar
  9. Browning, J.L., Seelig, J. 1980. Bilayers of phosphatidylserine: A deuterium and phosphorus nuclear magnetic resonance study.Biochemistry 19:1262–1270PubMedGoogle Scholar
  10. Chabre, M. 1975. X-ray diffraction studies of retinal rods: 1. Structure of the disc membrane, effect of illumination.Biochim. Biophys. Acta 382:322–335PubMedGoogle Scholar
  11. Chapman, D. 1975. Phase transitions and fluidity characteristics of lipids and cell membranes.Q. Rev. Biophys. 8:185–235Google Scholar
  12. Chen, Y.S., Hubbell, W.L. 1973. Temperature- and light-dependent structural changes in rhodopsin-lipid membranes.Exp. Eye Res. 17:517–532PubMedGoogle Scholar
  13. Cooper, A., Converse, C.A. 1976. Energetics of primary processes in visual excitation: Photocalorimetry of rhodopsin in rod outer segment membranes.Biochemistry 15:2970–2978PubMedGoogle Scholar
  14. Crain, R.C., Marinetti, G.V., O'Brien, D.F. 1978. Topology of amino phospholipids in bovine retinal rod outer segment disk membranes.Biochemistry 17:4186–4192PubMedGoogle Scholar
  15. Cullis, P.R., De Kruijff, B. 1978. The polymorphic phase behaviour of phosphatidylethanolamines of natural and synthetic origin. A31P NMR study.Biochim. Biophys. Acta 513:31–42PubMedGoogle Scholar
  16. Cullis, P.R., De Kruijff, B. 1979. Lipid polymorphism and the functional roles of lipids in biological membranes.Biochim. Biophys. Acta 559:399–420PubMedGoogle Scholar
  17. Cullis, P.R., Van Djick, P.W.M., De Kruijff, B., De Gier, J. 1978. Effects of cholesterol on the properties of equimolar mixtures of synthetic phosphatidylethanolamine and phosphatidylcholine. A31P NMR and differential scanning calorimetry study.Biochim. Biophys. Acta 513:21–30PubMedGoogle Scholar
  18. Cullis, P.R., Verkleij, A.J. 1979. Modulation of membrane structure by Ca2+ and dibucaine as detected by31P NMR.Biochim. Biophys. Acta 552:546–551PubMedGoogle Scholar
  19. Daemen, F.J.M. 1973. Vertebrate rod outer segment membranes.Biochim. Biophys. Acta 300:255–288PubMedGoogle Scholar
  20. Deese, A.J., Dratz, E.A., Brown, M.F. 1981a. Retinal rod outer segment lipids form bilayers in the presence and absence of rhodopsin: A31P NMP study.FEBS Lett. 124:93–99PubMedGoogle Scholar
  21. Deese, A.J., Dratz, E.A., Dahlquist, F.W., Paddy, M.R. 1981b. Interaction of rhodopsin with two unsaturated phosphatidylcholines: A deuterium nuclear magnetic resonance study.Biochemistry 20:6420–6427PubMedGoogle Scholar
  22. De Grip, W.J., Drenthe, E.H.S., Van Echteld, C.J.A., De Kruijff, B., Verkleij, A.J. 1979. A possible role of rhodopsin in maintaining bilayer structure in the photoreceptor membrane.Biochim. Biophys. Acta 558:330–337PubMedGoogle Scholar
  23. De Kruyff, B., Demel, R.A., Slotboom, A.J., Van Deenen, L.L.M., Rosenthal, A.F. 1973. The effect of the polar headgroup on the lipid-cholesterol interation: A monolayer and differential scanning calorimetry study.Biochim. Biophys. Acta 307:1–19PubMedGoogle Scholar
  24. Demel, R.A., Jansen, J.W.C.M., Van Dijck, P.W.M., Van Deenen, L.L.M. 1977. The preferential interaction of cholesterol with different classes of phospholipids.Biochim. Biophys. Acta 465:1–10PubMedGoogle Scholar
  25. Drenthe, E.H.S., Bonting, S.L., Daemen, F.J.M. 1980a. Transbilayer distribution of phospholipids in photoreceptor membrane studied with various phospholipases.Biochim. Biophys. Acta 603:117–129PubMedGoogle Scholar
  26. Drenthe, E.H.S., Klompmakers, A.A., Bonting, S.L., Daemen, F.J.M. 1980b. Transbilayer distribution of phospholipids in photoreceptor membrane studied with trinitrobenzenesulfonate alone and in combination with phospholipase D.Biochim. Biophys. Acta 603:130–141PubMedGoogle Scholar
  27. Estep, T.N., Mountcastle, D.B., Biltonen, R.L., Thompson, T.E. 1978. Studies on the anomalous thermotropic behavior of aqueous disperisons of dipalmitoylphosphatidylcholinecholesterol mixtures.Biochemistry 17:1984–1989PubMedGoogle Scholar
  28. Fisher, K.A. 1976. Analysis of membrane halves: cholesterol.Proc. Natl. Acad. Sci. USA 73:173–177PubMedGoogle Scholar
  29. Fung, B.K.-K., Hubbell, W.L. 1978. Organization of rhodopsin in photoreceptor membranes. 2. Transmembrane organization of bovine rhodopsin: Evidence from proteolysis and lactoperoxidase-catalyzed iodination of native and reconstituted membranes.Biochemistry 17:4403–4410PubMedGoogle Scholar
  30. Gally, H.U., Pluschke, G., Overath, P., Seelig, J. 1980. Structure ofEscherichia coli membranes. Fatty acyl chain order parameters of inner and outer membranes and derived liposomes.Biochemistry 19:1638–1643PubMedGoogle Scholar
  31. Gruner, S.M., Rothschild, K.J., Clark, N.A. 1982. X-ray diffraction and electron microscope study of phase separation in rod outer segment photoreceptor membrane multilayers.Biophys. J. 39:241–251PubMedGoogle Scholar
  32. Hendriks, T., Klompmakers, A.A., Daemen, F.J.M., Bonting, S.L. 1976. Biochemical aspects of the visual process: XXXII. Movement of sodium ions through bilayers composed of retinal and rod outer segment lipids.Biochim. Biophys. Acta 433:271–281Google Scholar
  33. Hong, K., Hubbell, W.L. 1973. Lipid requirements for rhodopsin regenerability.Biochemistry 12:4517–4523PubMedGoogle Scholar
  34. Hubbard, R. 1958. The thermal stability of rhodopsin and opsin.J. Gen. Physiol. 42:259–280PubMedGoogle Scholar
  35. Knudsen, P., Hubbell, W.L. 1978. Stability of rhodopsin in thetergent solutions.Membrane Biochem. 1:297–322Google Scholar
  36. Krebs, W., Kühn, H. 1977. Structure of isolated bovine rod outer segment membranes.Exp. Eye Res. 25:511–526Google Scholar
  37. Kusumi, A., Hyde, J.S. 1982. Spin-label sauration-transfer elecctron spin resonance detection of transient association of rhodopsin in reconstituted membranes.Biochemistry 21:5978–5983PubMedGoogle Scholar
  38. Lamola, A.A., Yamane, T., Zipp, A. 1974. Effects of detergents and high pressures upon the metarhodopsin I-metarhodopsin II equilibrium.Biochemistry 13:738–745PubMedGoogle Scholar
  39. Litman, B.J., Kalisky, O., Ottolenghi, M. 1981. Rhodopsinphospholipid interactions: Dependence of rat of the meta I to meta II transition on the level of associated disk phospholipid.Biochemistry 20:631–634PubMedGoogle Scholar
  40. Mabrey, S., Sturtevant, J.M. 1976. Investigation of phase transitions of lipids and lipid mixtures by high sensitivity defferential scanning calorimetry.Proc. Natl. Acad. Sci. USA 73:3862–3866PubMedGoogle Scholar
  41. Mabrey, S., Mateo, P.L., Sturtevant, J.M. 1978. High-sesitivity scanning calorimetric study of mixtures of cholesterol with dimyristoyl- and dipalmitoylphosphatidylcholines.Biochemistry 17:2464–2468PubMedGoogle Scholar
  42. Marsh, D., Watts, A., Pates, R.D., Uhl, R., Knowles, P.F., Esmann, M. 1982. ESR spin-label studies of lipid-protein interactions in membranes.Biophys. J. 37:265–274PubMedGoogle Scholar
  43. McCaslin, D.R., Tanford, C. 1981. Effects of detergent micelles on the recombination reaction of opsin and 11-cis-retinal.Biochemistry 20:5207–5212PubMedGoogle Scholar
  44. Miljanich, G.P., Mabrey, S.V., Brown, M.F., Sturtevant, J.M., Dratz, E.A. 1978. Differential scanning calorimetry of rod outer segment membranes and extracted phospholipids.Biophys. J. 21:135a Google Scholar
  45. Miljanich, G.P., Nemes, P.P., White, D.L., Dratz, E.A. 1981. The asymmetric transmembrane distribution of phosphatidylethanolamine, phosphatidylserine, and fatty acids of the bovine retinal rod outer segment disk membrane.J. Membrane Biol. 60:249–255Google Scholar
  46. Miljanich, G.P., Sklar, L.A., White, D.L., Dratz, E.A. 1979. Disaturated and dipolyunsaturated phospholipids in the bovine retinal rod outer segment disk membrane.Biochim. Biophys. Acta 522:294–306Google Scholar
  47. Mollevanger, L.C.P.J., DeGrip, W.J. 1984. Phase behavior of isolated photoreceptor membrane lipids is modulated by bivalent ions.FEBS Lett. 169:256–260PubMedGoogle Scholar
  48. Nemes, P.P., Miljanich, G.P., White, D.L., Dratz, E.A. 1980. Covalent modification of rhodopsin with imidoesters: Evidence for transmembrane arrangement of rhodopsin in rod outer segment disk membranes.Biochemistry 19:2067–2074PubMedGoogle Scholar
  49. O'Brien, D.F., Costa, L.F., Ott, R.A. 1977. Photochemiscal functionality of rhodopsin-phospholipid recombinant membranes.Biochemistry 16:1295–1303PubMedGoogle Scholar
  50. Op den Kamp, J.A.F. 1979. Lipid asymmetry in membranesAnnu. Rev. Biochem. 48:47–71PubMedGoogle Scholar
  51. Phillips, M.C., Ladbrooke, B.D., Chapman, D. 1970. Molecular interactions in mixed lecithin systems.Biochim. Biophys. Acta 196:35–44PubMedGoogle Scholar
  52. Raubach, R.A., Franklin, L.K., Dratz, E.A. 1974a. A rapid method for the purification of rod outer segment disk membranes.Vision Res. 14:335–337PubMedGoogle Scholar
  53. Raubach, R.A., Nemes, P.P., Dratz, E.A. 1974b. Chemical labeling and freeze-fracture studies on the localization of rhodopsin in the rod outer segment disk membrane.Exp. Eye Res. 18:1–12PubMedGoogle Scholar
  54. Rothschild, K.J., Andrew, J.R., De Grip, W.J., Stanley, H.E. 1976. Opsin structure probed by Raman spectroscopy of photoreceptor membranes.Science 191:1176–1178PubMedGoogle Scholar
  55. Sklar, L.A., Dratz, E.A. 1980. Analysis of membrane bilayer asymmetry using parinatric acid fluorescent probes.FEBS Lett. 118:308–310PubMedGoogle Scholar
  56. Sklar, L.A., Miljanich, G.P., Bursten, S.L., Dratz, E.A. 1979a. Thermal lateral phase separations in bovine retinal rod outer segment membranes and phospholipids as evidenced by parinaric acid fluorescence polarization and energy transfer.J. Biol. Chem. 254:9583–9591PubMedGoogle Scholar
  57. Sklar, L.A., Miljanich, G.P., Dratz, E.A. 1979b. Phospholipid lateral phase separation and the partition ofcis-parinaric acid andtrans-parinaric acid among aqueous, solid lipid, and fluid lipid phases.Biochemistry 18:1707–1716PubMedGoogle Scholar
  58. Smith, H.G., Fager, R.S., Litman, B.J. 1977. Light-activated calcium release from sonicated bovine retinal rod outer segment disks.Biochemistry 16:1399–1405PubMedGoogle Scholar
  59. Stone, W.L., Farnsworth, C.C., Dratz, E.A. 1979. Reinvestigation of the faty acid content of bovine, rat and frog retinal rod outer segments.Exp. Eye Res 28:387–397PubMedGoogle Scholar
  60. Stubbs, G.W., Litman, B.J., Barenholz, Y. 1976a. Microviscosity of the hydrocarbon region of the bovine rod outer segment disk membrane determined by fluorescent probe measurements.Biochemistry 15:2766–2772PubMedGoogle Scholar
  61. Stubbs, G.W., Litman, B.J. 1978. Effect of alterations in the amphipathic microenvironment on the conformational stability of bovine opsin. I. Mechanism of solubilization of disk membranes by the nonionic detergent, octyl glucoside.Biochemistry 17:215–219PubMedGoogle Scholar
  62. Stubbs, G.W., Smith, H.G., Litman, B.J. 1976b. Alkyl glucosides as effective solubilizing agents for bovine rhodopsin. A comparison with several commonly used detergents.Biochim. Biophy. Acta 425:46–56Google Scholar
  63. Van Dijck, P.W.M., De Kruijff, B., Van Deenen, L.L.M., De Gier, J., Demel, R.A. 1976. The preference of cholesterol for phosphatidylcholine in mixed phosphatidylcholine-phosphatidylethanolamine bilayers.Biochim. Biophys. Acta 455:576–587PubMedGoogle Scholar
  64. Watts, A., Davoust, J., Marsh, D., Devaux, P.F. 1981. Distinct states of lipid mobility in bovine rod outer segment membranes. Resolution of spin label results.Biochim. Biophys. Acta 643:673–676PubMedGoogle Scholar
  65. Watts, A., Volotovski, I.D., Marsh, D. 1979. Rhodopsin-lipid associations in bovine rod outer segment membranes. Identification of immobilized lipid by spin-labels.Biochemistry 18:5006–5013PubMedGoogle Scholar
  66. Wilkinson, D.A., Nagle, J.F. 1981. Dilatometry and calorimetry of saturated phosphatidylethanolamine dispersions.Biochemistry 20:187–192PubMedGoogle Scholar

Copyright information

© Springer-Verlag 1985

Authors and Affiliations

  • George P. Miljanich
    • 1
  • Michael F. Brown
    • 2
  • Susan Mabrey-Gaud
    • 3
  • Edward A. Dratz
    • 1
  • Julian M. Sturtevant
    • 3
  1. 1.Chemistry Board of Studies, Division of Natural SciencesUniversity of CaliforniaSanta Cruz
  2. 2.Department of Chemistry and Biophysics ProgramUniversity of VirginiaCharlottesville
  3. 3.Department of ChemistryYale UniversityNew Haven

Personalised recommendations