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. 2001 Dec;81(6):3346–3362. doi: 10.1016/S0006-3495(01)75968-1

Polylysine-induced 2H NMR-observable domains in phosphatidylserine/phosphatidylcholine lipid bilayers.

C M Franzin 1, P M Macdonald 1
PMCID: PMC1301792  PMID: 11720998

Abstract

The interaction of three polylysines, Lys(5) (N = 5), Lys(30) (N = 30), and Lys(100) (N = 100), where N is the number of lysine residues per chain, with phosphatidylserine-containing lipid bilayer membranes was investigated using 2H NMR spectroscopy. Lys(30) and Lys(100) added to multilamellar vesicles composed of (70:30) (mol:mol) mixtures of choline-deuterated 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) + 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoserine (POPS) produced two resolvable 2H NMR spectral components under conditions of low ionic strength and for cases where the global anionic lipid charge was in excess over the global cationic polypeptide charge. The intensities and quadrupolar splittings of the two spectral components were consistent with the existence of polylysine-bound domains enriched in POPS, in coexistence with polylysine-free domains depleted in POPS. Lys(5), however, yielded no 2H NMR resolvable domains. Increasing ionic strength caused domains to become diffuse and eventually dissipate entirely. At physiological salt concentrations, only Lys(100) yielded 2H NMR-resolvable domains. Therefore, under physiological conditions of ionic strength, pH, and anionic lipid bilayer content, and in the absence of other, e.g., hydrophobic, contributions to the binding free energy, the minimum number of lysine residues sufficient to produce spectroscopically resolvable POPS-enriched domains on the 2H NMR millisecond timescale may be fewer than 100, but is certainly greater than 30.

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Selected References

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  1. Ben-Tal N., Honig B., Peitzsch R. M., Denisov G., McLaughlin S. Binding of small basic peptides to membranes containing acidic lipids: theoretical models and experimental results. Biophys J. 1996 Aug;71(2):561–575. doi: 10.1016/S0006-3495(96)79280-9. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Beschiaschvili G., Seelig J. Melittin binding to mixed phosphatidylglycerol/phosphatidylcholine membranes. Biochemistry. 1990 Jan 9;29(1):52–58. doi: 10.1021/bi00453a007. [DOI] [PubMed] [Google Scholar]
  3. Beswick V., Roux M., Navarre C., Coïc Y. M., Huynh-Dinh T., Goffeau A., Sanson A., Neumann J. M. 1H- and 2H-NMR studies of a fragment of PMP1, a regulatory subunit associated with the yeast plasma membrane H(+)-ATPase. Conformational properties and lipid-peptide interactions. Biochimie. 1998 May-Jun;80(5-6):451–459. doi: 10.1016/s0300-9084(00)80012-7. [DOI] [PubMed] [Google Scholar]
  4. Carnegie P. R. Amino acid sequence of the encephalitogenic basic protein from human myelin. Biochem J. 1971 Jun;123(1):57–67. doi: 10.1042/bj1230057. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Carrier D., Mantsch H. H., Wong P. T. Protective effect of lipidic surfaces against pressure-induced conformational changes of poly(L-lysine). Biochemistry. 1990 Jan 9;29(1):254–258. doi: 10.1021/bi00453a034. [DOI] [PubMed] [Google Scholar]
  6. Carrier D., Pézolet M. Investigation of polylysine-dipalmitoylphosphatidylglycerol interactions in model membranes. Biochemistry. 1986 Jul 15;25(14):4167–4174. doi: 10.1021/bi00362a027. [DOI] [PubMed] [Google Scholar]
  7. Cevc G. Membrane electrostatics. Biochim Biophys Acta. 1990 Oct 8;1031(3):311–382. doi: 10.1016/0304-4157(90)90015-5. [DOI] [PubMed] [Google Scholar]
  8. Collins K. D. Charge density-dependent strength of hydration and biological structure. Biophys J. 1997 Jan;72(1):65–76. doi: 10.1016/S0006-3495(97)78647-8. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Cullis P. R., de Kruijff B. Lipid polymorphism and the functional roles of lipids in biological membranes. Biochim Biophys Acta. 1979 Dec 20;559(4):399–420. doi: 10.1016/0304-4157(79)90012-1. [DOI] [PubMed] [Google Scholar]
  10. Davenport L., Knutson J. R., Brand L. Fluorescence studies of membrane dynamics and heterogeneity. Subcell Biochem. 1989;14:145–188. doi: 10.1007/978-1-4613-9362-7_4. [DOI] [PubMed] [Google Scholar]
  11. Denisov G., Wanaski S., Luan P., Glaser M., McLaughlin S. Binding of basic peptides to membranes produces lateral domains enriched in the acidic lipids phosphatidylserine and phosphatidylinositol 4,5-bisphosphate: an electrostatic model and experimental results. Biophys J. 1998 Feb;74(2 Pt 1):731–744. doi: 10.1016/S0006-3495(98)73998-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Eylar E. H., Brostoff S., Hashim G., Caccam J., Burnett P. Basic A1 protein of the myelin membrane. The complete amino acid sequence. J Biol Chem. 1971 Sep 25;246(18):5770–5784. [PubMed] [Google Scholar]
  13. Glaser M., Wanaski S., Buser C. A., Boguslavsky V., Rashidzada W., Morris A., Rebecchi M., Scarlata S. F., Runnels L. W., Prestwich G. D. Myristoylated alanine-rich C kinase substrate (MARCKS) produces reversible inhibition of phospholipase C by sequestering phosphatidylinositol 4,5-bisphosphate in lateral domains. J Biol Chem. 1996 Oct 18;271(42):26187–26193. doi: 10.1074/jbc.271.42.26187. [DOI] [PubMed] [Google Scholar]
  14. Gliss C., Clausen-Schaumann H., Günther R., Odenbach S., Randl O., Bayerl T. M. Direct detection of domains in phospholipid bilayers by grazing incidence diffraction of neutrons and atomic force microscopy. Biophys J. 1998 May;74(5):2443–2450. doi: 10.1016/S0006-3495(98)77952-4. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Hammes G. G., Schullery S. E. Structure of macromolecular aggregates. II. Construction of model membranes from phospholipids and polypeptides. Biochemistry. 1970 Jun 23;9(13):2555–2563. doi: 10.1021/bi00815a001. [DOI] [PubMed] [Google Scholar]
  16. Hartmann W., Galla H. J. Binding of polylysine to charged bilayer membranes: molecular organization of a lipid.peptide complex. Biochim Biophys Acta. 1978 Jun 2;509(3):474–490. doi: 10.1016/0005-2736(78)90241-9. [DOI] [PubMed] [Google Scholar]
  17. Huang J., Feigenson G. W. Monte Carlo simulation of lipid mixtures: finding phase separation. Biophys J. 1993 Nov;65(5):1788–1794. doi: 10.1016/S0006-3495(93)81234-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Huang J., Swanson J. E., Dibble A. R., Hinderliter A. K., Feigenson G. W. Nonideal mixing of phosphatidylserine and phosphatidylcholine in the fluid lamellar phase. Biophys J. 1993 Feb;64(2):413–425. doi: 10.1016/S0006-3495(93)81382-1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Jovin T. M., Vaz W. L. Rotational and translational diffusion in membranes measured by fluorescence and phosphorescence methods. Methods Enzymol. 1989;172:471–513. doi: 10.1016/s0076-6879(89)72030-9. [DOI] [PubMed] [Google Scholar]
  20. Kim J., Mosior M., Chung L. A., Wu H., McLaughlin S. Binding of peptides with basic residues to membranes containing acidic phospholipids. Biophys J. 1991 Jul;60(1):135–148. doi: 10.1016/S0006-3495(91)82037-9. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Kim J., Shishido T., Jiang X., Aderem A., McLaughlin S. Phosphorylation, high ionic strength, and calmodulin reverse the binding of MARCKS to phospholipid vesicles. J Biol Chem. 1994 Nov 11;269(45):28214–28219. [PubMed] [Google Scholar]
  22. Kinnunen P. K., Kõiv A., Lehtonen J. Y., Rytömaa M., Mustonen P. Lipid dynamics and peripheral interactions of proteins with membrane surfaces. Chem Phys Lipids. 1994 Sep 6;73(1-2):181–207. doi: 10.1016/0009-3084(94)90181-3. [DOI] [PubMed] [Google Scholar]
  23. Kleinschmidt J. H., Marsh D. Spin-label electron spin resonance studies on the interactions of lysine peptides with phospholipid membranes. Biophys J. 1997 Nov;73(5):2546–2555. doi: 10.1016/S0006-3495(97)78283-3. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Köchy T, Bayerl TM. Lateral diffusion coefficients of phospholipids in spherical bilayers on a solid support measured by 2H-nuclear-magnetic-resonance relaxation. Phys Rev E Stat Phys Plasmas Fluids Relat Interdiscip Topics. 1993 Mar;47(3):2109–2116. doi: 10.1103/physreve.47.2109. [DOI] [PubMed] [Google Scholar]
  25. Laroche G., Carrier D., Pézolet M. Study of the effect of poly(L-lysine) on phosphatidic acid and phosphatidylcholine/phosphatidic acid bilayers by raman spectroscopy. Biochemistry. 1988 Aug 23;27(17):6220–6228. doi: 10.1021/bi00417a005. [DOI] [PubMed] [Google Scholar]
  26. Laroche G., Dufourc E. J., Pézolet M., Dufourcq J. Coupled changes between lipid order and polypeptide conformation at the membrane surface. A 2H NMR and Raman study of polylysine-phosphatidic acid systems. Biochemistry. 1990 Jul 10;29(27):6460–6465. doi: 10.1021/bi00479a018. [DOI] [PubMed] [Google Scholar]
  27. Luan P., Yang L., Glaser M. Formation of membrane domains created during the budding of vesicular stomatitis virus. A model for selective lipid and protein sorting in biological membranes. Biochemistry. 1995 Aug 8;34(31):9874–9883. doi: 10.1021/bi00031a008. [DOI] [PubMed] [Google Scholar]
  28. Macdonald P. M., Crowell K. J., Franzin C. M., Mitrakos P., Semchyschyn D. J. Polyelectrolyte-induced domains in lipid bilayer membranes: the deuterium NMR perspective. Biochem Cell Biol. 1998;76(2-3):452–464. doi: 10.1139/bcb-76-2-3-452. [DOI] [PubMed] [Google Scholar]
  29. Macdonald P. M., Crowell K. J., Franzin C. M., Mitrakos P., Semchyschyn D. 2H NMR and polyelectrolyte-induced domains in lipid bilayers. Solid State Nucl Magn Reson. 2000 May;16(1-2):21–36. doi: 10.1016/s0926-2040(00)00051-5. [DOI] [PubMed] [Google Scholar]
  30. Marassi F. M., Macdonald P. M. Response of the phosphatidylcholine headgroup to membrane surface charge in ternary mixtures of neutral, cationic, and anionic lipids: a deuterium NMR study. Biochemistry. 1992 Oct 20;31(41):10031–10036. doi: 10.1021/bi00156a024. [DOI] [PubMed] [Google Scholar]
  31. Marassi F. M., Shivers R. R., Macdonald P. M. Resolving the two monolayers of a lipid bilayer in giant unilamellar vesicles using deuterium nuclear magnetic resonance. Biochemistry. 1993 Sep 28;32(38):9936–9943. doi: 10.1021/bi00089a009. [DOI] [PubMed] [Google Scholar]
  32. May S., Harries D., Ben-Shaul A. Lipid demixing and protein-protein interactions in the adsorption of charged proteins on mixed membranes. Biophys J. 2000 Oct;79(4):1747–1760. doi: 10.1016/S0006-3495(00)76427-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. McLaughlin S., Aderem A. The myristoyl-electrostatic switch: a modulator of reversible protein-membrane interactions. Trends Biochem Sci. 1995 Jul;20(7):272–276. doi: 10.1016/s0968-0004(00)89042-8. [DOI] [PubMed] [Google Scholar]
  34. Mendelsohn R., Moore D. J. Vibrational spectroscopic studies of lipid domains in biomembranes and model systems. Chem Phys Lipids. 1998 Nov;96(1-2):141–157. doi: 10.1016/s0009-3084(98)00085-1. [DOI] [PubMed] [Google Scholar]
  35. Mitrakos P., Macdonald P. M. Cationic amphiphile interactions with polyadenylic acid as probed via 2H-NMR. Biochim Biophys Acta. 1998 Sep 23;1374(1-2):21–33. doi: 10.1016/s0005-2736(98)00128-x. [DOI] [PubMed] [Google Scholar]
  36. Mitrakos P., Macdonald P. M. DNA-induced lateral segregation of cationic amphiphiles in lipid bilayer membranes as detected via 2H NMR. Biochemistry. 1996 Dec 24;35(51):16714–16722. doi: 10.1021/bi961911h. [DOI] [PubMed] [Google Scholar]
  37. Mitrakos P., Macdonald P. M. Domains in cationic lipid plus polyelectrolyte bilayer membranes: detection and characterization via 2H nuclear magnetic resonance. Biochemistry. 1997 Nov 4;36(44):13646–13656. doi: 10.1021/bi971324b. [DOI] [PubMed] [Google Scholar]
  38. Mitrakos P., Macdonald P. M. Polyelectrolyte molecular weight and electrostatically-induced domains in lipid bilayer membranes. Biomacromolecules. 2000 Fall;1(3):365–376. doi: 10.1021/bm000029v. [DOI] [PubMed] [Google Scholar]
  39. Monette M., Lafleur M. Modulation of melittin-induced lysis by surface charge density of membranes. Biophys J. 1995 Jan;68(1):187–195. doi: 10.1016/S0006-3495(95)80174-8. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Mosior M., McLaughlin S. Peptides that mimic the pseudosubstrate region of protein kinase C bind to acidic lipids in membranes. Biophys J. 1991 Jul;60(1):149–159. doi: 10.1016/S0006-3495(91)82038-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. 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]
  42. Raudino A. Lateral inhomogeneous lipid membranes: theoretical aspects. Adv Colloid Interface Sci. 1995 May 30;57:229–285. doi: 10.1016/0001-8686(95)00243-j. [DOI] [PubMed] [Google Scholar]
  43. Roux M., Beswick V., Coïc Y. M., Huynh-Dinh T., Sanson A., Neumann J. M. PMP1 18-38, a yeast plasma membrane protein fragment, binds phosphatidylserine from bilayer mixtures with phosphatidylcholine: a (2)H-NMR study. Biophys J. 2000 Nov;79(5):2624–2631. doi: 10.1016/S0006-3495(00)76501-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Roux M., Neumann J. M., Bloom M., Devaux P. F. 2H and 31P NMR study of pentalysine interaction with headgroup deuterated phosphatidylcholine and phosphatidylserine. Eur Biophys J. 1988;16(5):267–273. doi: 10.1007/BF00254062. [DOI] [PubMed] [Google Scholar]
  45. Roux M., Neumann J. M., Hodges R. S., Devaux P. F., Bloom M. Conformational changes of phospholipid headgroups induced by a cationic integral membrane peptide as seen by deuterium magnetic resonance. Biochemistry. 1989 Mar 7;28(5):2313–2321. doi: 10.1021/bi00431a050. [DOI] [PubMed] [Google Scholar]
  46. Rydall J. R., Macdonald P. M. Investigation of anion binding to neutral lipid membranes using 2H NMR. Biochemistry. 1992 Feb 4;31(4):1092–1099. doi: 10.1021/bi00119a018. [DOI] [PubMed] [Google Scholar]
  47. Scherer P. G., Seelig J. Electric charge effects on phospholipid headgroups. Phosphatidylcholine in mixtures with cationic and anionic amphiphiles. Biochemistry. 1989 Sep 19;28(19):7720–7728. doi: 10.1021/bi00445a030. [DOI] [PubMed] [Google Scholar]
  48. 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]
  49. Seelig J., Macdonald P. M., Scherer P. G. Phospholipid head groups as sensors of electric charge in membranes. Biochemistry. 1987 Dec 1;26(24):7535–7541. doi: 10.1021/bi00398a001. [DOI] [PubMed] [Google Scholar]
  50. Stillwell W., Jenski L. J., Zerouga M., Dumaual A. C. Detection of lipid domains in docasahexaenoic acid-rich bilayers by acyl chain-specific FRET probes. Chem Phys Lipids. 2000 Feb;104(2):113–132. doi: 10.1016/s0009-3084(99)00122-x. [DOI] [PubMed] [Google Scholar]
  51. Swierczynski S. L., Blackshear P. J. Membrane association of the myristoylated alanine-rich C kinase substrate (MARCKS) protein. Mutational analysis provides evidence for complex interactions. J Biol Chem. 1995 Jun 2;270(22):13436–13445. doi: 10.1074/jbc.270.22.13436. [DOI] [PubMed] [Google Scholar]
  52. Takahashi H., Yasue T., Ohki K., Hatta I. Structure and phase behaviour of dimyristoylphosphatidic acid/poly(L-lysine) systems. Mol Membr Biol. 1996 Oct-Dec;13(4):233–240. doi: 10.3109/09687689609160601. [DOI] [PubMed] [Google Scholar]
  53. Tamm L. K., Seelig J. Lipid solvation of cytochrome c oxidase. Deuterium, nitrogen-14, and phosphorus-31 nuclear magnetic resonance studies on the phosphocholine head group and on cis-unsaturated fatty acyl chains. Biochemistry. 1983 Mar 15;22(6):1474–1483. doi: 10.1021/bi00275a023. [DOI] [PubMed] [Google Scholar]
  54. Taniguchi H., Manenti S. Interaction of myristoylated alanine-rich protein kinase C substrate (MARCKS) with membrane phospholipids. J Biol Chem. 1993 May 15;268(14):9960–9963. [PubMed] [Google Scholar]
  55. Tocanne J. F., Cézanne L., Lopez A., Piknova B., Schram V., Tournier J. F., Welby M. Lipid domains and lipid/protein interactions in biological membranes. Chem Phys Lipids. 1994 Sep 6;73(1-2):139–158. doi: 10.1016/0009-3084(94)90179-1. [DOI] [PubMed] [Google Scholar]
  56. Tsui F. C., Ojcius D. M., Hubbell W. L. The intrinsic pKa values for phosphatidylserine and phosphatidylethanolamine in phosphatidylcholine host bilayers. Biophys J. 1986 Feb;49(2):459–468. doi: 10.1016/S0006-3495(86)83655-4. [DOI] [PMC free article] [PubMed] [Google Scholar]
  57. Victor K., Jacob J., Cafiso D. S. Interactions controlling the membrane binding of basic protein domains: phenylalanine and the attachment of the myristoylated alanine-rich C-kinase substrate protein to interfaces. Biochemistry. 1999 Sep 28;38(39):12527–12536. doi: 10.1021/bi990847b. [DOI] [PubMed] [Google Scholar]
  58. Wada A. The alpha-helix as an electric macro-dipole. Adv Biophys. 1976:1–63. [PubMed] [Google Scholar]
  59. Wohlgemuth R., Waespe-Sarcevic N., Seelig J. Bilayers of phosphatidylglycerol. A deuterium and phosphorus nuclear magnetic resonance study of the head-group region. Biochemistry. 1980 Jul 8;19(14):3315–3321. doi: 10.1021/bi00555a033. [DOI] [PubMed] [Google Scholar]
  60. Zachowski A., Devaux P. F. Non-uniform distribution of phospholipids in (Na+ + K+)-ATPase-rich membranes from Torpedo marmorata electric organ evidenced by spin-spin interactions between spin-labeled phospholipids. FEBS Lett. 1983 Nov 14;163(2):245–249. doi: 10.1016/0014-5793(83)80828-x. [DOI] [PubMed] [Google Scholar]
  61. de Kruijff B., Rietveld A., Telders N., Vaandrager B. Molecular aspects of the bilayer stabilization induced by poly(L-lysines) of varying size in cardiolipin liposomes. Biochim Biophys Acta. 1985 Nov 7;820(2):295–304. doi: 10.1016/0005-2736(85)90124-5. [DOI] [PubMed] [Google Scholar]

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