Skip to main content
SpringerLink
Log in

Influence of obstacles on lipid lateral diffusion: computer simulation of FRAP experiments and application to proteoliposomes and biomembranes

  • Published:
European Biophysics Journal Aims and scope Submit manuscript

Abstract

Fluorescence Recovery After Photobleaching experiments were simulated using a computer approach in which a membrane lipid leaflet was mimicked using a triangular lattice obstructed with randomly distributed immobile and non-overlapping circular obstacles. Influence of the radius r and area fraction c of these obstacles and of the radius R of the observation area on the relative diffusion coefficient D * (Eq. (1)) and mobile fraction M was analyzed. A phenomenological equation relating D * to r and c was established. Fitting this equation to the FRAP data we obtained with the probe NBD-PC embedded in bacteriorhodopsin/egg-PC multilayers suggests that this transmembrane protein rigidifies the surrounding lipid phase over a distance of about 18 Å (≅two lipid layers) from the protein surface. In contrast, analysis of published diffusion constants obtained for lipids in the presence of gramicidin suggests that in terms of lateral diffusion, this relatively small polypeptide does not significantly affect the surrounding lipid phase. With respect to the mobile fraction M, and for point obstacles above the percolation threshold, an increase in R led to a decrease in M which can be associated with the existence of closed domains whose average size and diffusion properties can be determined. Adaptation of this model to the re-interpretation of the FRAP data obtained by Yechiel and Edidin (J Cell Biol (1987) 115:755–760) for the plasma membrane of human fibroblasts consistently leads to the suggestion that the lateral organization of this membrane would be of the confined type, with closed lipid domains of ≅0.5 µm2 in area.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

Abbreviations

BR:

bacteriorhodopsin

DMPC:

dimyristoylphosphatidylcholine

diOC18:

dioctadecyloxatricarbocyanine

egf-PC:

egg-yolk phosphatidylcholine

NBD-PC:

1-acyl2-[t2-[(7-nitro-2-1,3-benzoxadiazol-4-yl)amino]dodecanoyl]-sn-glycero-3-phosphocholine

MOPS:

3-[N-morpholino]propane sulfonic acid

FRAP:

Fluoresence Recovery After photobleaching

D:

observed diffusion coefficient

D0:

diffusion coefficient in the absence of obstacles

D * :

relative diffusion constant (Eq. 1)

M :

mobile fraction

c :

obstacle area fraction

r :

obstacle radius

R :

observation area radius

r d :

diffusion area radius

References

  • Abney JR, Scalettar BA, Owicki JC (1989) Self diffusion of interacting membrane proteins. Biophys J 55:817–833

    Google Scholar 

  • Almeida PFF, Vaz WLC, Thompson TE (1992) Lateral diffusion and percolation in two-phase, two-component lipid bilayers. Topology of the solid-phase domains in plane and across the lipid bilayer. Biochemistry 31:7198–7210

    Google Scholar 

  • Axelrod D, Koppel DE, Schlessinger J, Elson E, Webb WW (1976) Mobility measurement by analysis of fluorescence photobleaching recovery kinetics. Biophys J 16:1055–1069

    Google Scholar 

  • Blackwell ME, Whitmarsh J (1990) Effect of integral membrane proteins on the lateral mobility of plastoquinone in phosphatidylcholine proteoliposomes. Biophys J 58: 1259–1271

    Google Scholar 

  • Blaurock AE (1975) Bacteriorhodopsin: a trans-membrane pump containing alpha-helix. J Mol Biol 93:139–158

    Google Scholar 

  • Bloom M, Evans E, Mouritsen OG (1991) Physical properties of the fluid lipid-bilayer component of cell membranes: A perspective. Q Rev Biophys 24:293–397

    Google Scholar 

  • Bussell SJ, Hammer DA, Koch DL (1994) The effect of hydrodynamic interactions on the tracer and gradient diffusion of integral membrane proteins in lipid bilayers. J Fluid Mech 258: 167–190

    Google Scholar 

  • Cherry RJ (1979) Rotational and lateral diffusion of membrane proteins. Biochim Biophys Acta 559:289–327

    Google Scholar 

  • Edidin M (1987) Rotational and lateral diffusion of membrane proteins and lipids: phenomena and function. Curr Top Membr Transp 29:91–127

    Google Scholar 

  • Eisinger J, Flores J, Petersen WP (1986) A Milling crowd model for local and long range obstructed lateral diffusion. Mobility of excimeric probes in the membrane of intact erythrocytes. Biophys J 49:987–1001

    Google Scholar 

  • Fery-Forgues S, Fayet JP, Lopez A (1993) Drastic changes in the fluorescence properties of NBD probes with the polarity of the medium: involvement of a TICT state. J Photochem Photobiol A: Chem 70:229–243

    Google Scholar 

  • Gulik-Krzywicki T, Seigneuret M, Rigaud JL (1987) Monomer-oligomer equilibrium of bacteriorhodopsin in reconstituted proteoliposomes. A freeze-fracture electron microscopy study. J Biol Chem 262:15580–15588

    Google Scholar 

  • Henderson R, Unwin PNT (1975) Three-dimensional model of purple membrane obtained by electron microscopy. Nature 257: 28–32

    Google Scholar 

  • Heyn MP, Dencher NA (1982) Reconstitution of monomeric bacteriorhodopsin into phospholipid vesicles. Methods Enzymol 88: 31–35

    Google Scholar 

  • Heyn MP, Blume A, Rehorek M, Dencher NA (1981) Calorimetric and fluorescence depolarization studies on the lipid phase transition of bacteriorhodopsin-dimyristoylphosphatidylcholine vesicles. Biochemistry 20:7109–7115

    Google Scholar 

  • Jähnig F (1981) Critical effects from lipid-protein interaction in membranes. I. Theoretical description. Biophys J 36:329–345

    Google Scholar 

  • Kusumi A (1994) Confined lateral diffusion of membrane receptors as studied by single particle tracking and laser tweezers. Biophys J 66: A18

    Google Scholar 

  • Lentz BR (1988) Organization of membrane lipids by intrinsic membrane proteins. In: Advances in membrane fluidity. Vol. 2. Aloia RC, Curtain CC, Goron LM (eds) A. R. Liss, New York, pp 141–161

    Google Scholar 

  • Lewis BA, Engelman DM (1983) Bacteriorhodopsin remains dispersed in fluid phospholipid bilayers over a wide range of bilayer thicknesses. J Mol Biol 166: 203–211

    Google Scholar 

  • Lopez A, Dupou L, Altibelli A, Trotard J, Tocanne JF (1988) Fluorescence Recovery After Photobleaching (FRAP) experiments under conditions of uniform disk illumination: Critical comparison of analytical solutions, and a new mathematical method for calculation of diffusion coefficient D. Biophys J 53:963–970

    Google Scholar 

  • Marsh D, Watts A (1988) Association of lipids with membrane proteins. In: Advances in membrane fluidity. Vol. 2. Aloia RC, Curtain CC, Gordon LM (eds) A. R. Liss, New York, pp 163–200

    Google Scholar 

  • Oesterhelt D, Stoeckenius W (1974) Isolation of the cell membrane of Halobacterium halobium and its fractionation into red and purple membrane. Methods Enzymol 31:667–678

    Google Scholar 

  • Onoda GY, Liniger EG (1986) Experimental determination of the random-parking limit in two dimensions. Phys Rev A 33: 715–716

    Google Scholar 

  • Owicki JC, Springgate MW, McConnell HM (1978) Theoretical study of protein-lipid interactions in bilayer membranes. Proc Natl Acad Sci, USA 75:1616–1619

    Google Scholar 

  • Peschke J, Riegler J, Möhwald H (1987) Quantitative analysis of membrane distorsions induced by mismatch of protein and lipid hydrophobic thickness. Eur Biophys J 14:385–391

    Google Scholar 

  • Peters R, Cherry RJ (1982) Lateral and rotational diffusion of bacteriorhodopsin in lipid bilayers: Experimental test of the Saffman-Delbrück equations. Proc Natl Acad Sci, USA 79: 4317–4321

    Google Scholar 

  • Piknova B, Perochon E, Tocanne JF (1993) Hydrophobic mismatch and long-range protein-lipid interactions in bacteriorhodopsin/phosphatidylcholine vesicles. Eur J Biochem 218:385–396

    Google Scholar 

  • Pink DA (1985) Protein lateral movement in lipid bilayers. Simulation studies of its dependence upon protein concentration. Biochim Biophys Acta 863:9–17

    Google Scholar 

  • Rehorek M, Dencher NA, Heyn MP (1985) Long-range lipid-protein interactions. Evidence from time-resolved fluorescence depolarization and energy-transfer experiments with bacteriorhodopsin-dimyristoylphosphatidylcholine vesicles. Biochemistry 24:5980–5988

    Google Scholar 

  • Saxton MJ (1982) Lateral diffusion in an archipelago. Effects of impermeable patches on diffusion in a cell membrane. Biophys J 39:165–173

    Google Scholar 

  • Saxton MJ (1987) Lateral diffusion in an archipelago: the effect of mobile obstacles. Biophys J 52:989–997

    Google Scholar 

  • Saxton MJ (1989a) The spectrin network as a barrier to lateral diffusion in erythrocytes. A percolation analysis. Biophys 155: 21–28

    Google Scholar 

  • Saxton MJ (1989b) Lateral diffusion in an archipelago: distance dependence of the diffusion coefficient. Biophys J 56:615–622

    Google Scholar 

  • Saxton MJ (1992) Lateral diffusion and aggregation. A Monte Carlo study. Biophys J 61:119–128

    Google Scholar 

  • Scalettar BA, Abney JR (1991) Molecular crowding and protein diffusion in biological membranes. Comments Mol Cell Biophys 7:79–107

    Google Scholar 

  • Scotto AW, Zakim D (1988) Reconstitution of membrane proteins. Spontaneous incorporation of integral membrane proteins into preformed bilayers of pure phospholipid. J Biol Chem 34: 18500–18506

    Google Scholar 

  • Sperotto MM, Mouritsen OG (1988) Dependence of lipid membrane phase transition temperature on the mismatch of protein and lipid hydrophobic thickness. Eur Biophys J 16:1–10

    Google Scholar 

  • Sperotto MM, Mouritsen OG (1991) Monte Carlo simulation studies of lipid order parameter profiles near integral membrane proteins. Biophys J 59:261–270

    Google Scholar 

  • Stauffer D, Abarony A (1992) Introduction to percolation theory. Taylor and Francis, London, p 124

    Google Scholar 

  • Tank DW, Wu ES, Meets PR, Webb WW (1982) Lateral diffusion of gramicidin C in phospholipid multibilayers. Effects of cholesterol and high gramicidin concentration. Biophys 140: 129–135

    Google Scholar 

  • Thomas JL, Holowka D, Baird B, Webb W (1994) Large-scale co-aggregation of fluorescent lipid probes with cell surface proteins. J Cell Biol 125:795–802

    Google Scholar 

  • Tocanne JF (1992) Detection of lipid domains in biological membranes. Comments Mol Cell Biophys 8:53–72

    Google Scholar 

  • Tocanne JF, Dupou-Cezanne L, Lopez A (1994a) Lateral diffusion of lipids in model and natural membranes. Prog Lipid Res 33:203–237

    Google Scholar 

  • Tocanne JF, Cézanne L, Lopez A, Perochon E, Piknova B, Schram V, Tournier JF, Welby M (1994b) Lipid domains and lipid/protein interactions in biological membranes. Chem Phys Lipids (in press)

  • Vaz WLC, Derzko ZI, Jacobson KA (1982) Photobleaching measurements of the lateral diffusion of lipids and proteins in artificial phospholipid bilayer membranes. In: Membrane reconstitution. Poste et Nicholson (eds) Elsevier Biomedical Press, pp 83–136

  • Vaz WLC, Melo ECC, Thompson TE (1989) Translational diffusion and fluid domains connectivity in a two-component, two-phase phospholipid bilayer. Biophys J 56:869–876

    Google Scholar 

  • Yeagle PL (1988) Use of NMR to study lipid phase states and protein-induced lipid domains. In: Advances in membrane fluidity. Vol. 1. Aloia RC, Curtain CC, Gordon LM (eds) A. R. Liss, New York, pp 67–295

    Google Scholar 

  • Yechiel E, Edidin M (1987) Micrometer-scale domains in fibroblast plasma membranes. J Cell Biol 115:755–760

    Google Scholar 

  • Wallace BA, Ravikumar K (1992) The gramicidin pore: Crystal structure of a cesium complex. Science 241:182–187

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Laboratoire de Pharmacologie et Toxicologie Fondamentales du CNRS (Département III), 118 route de Narbonne, F-31062, Toulouse Cedex, France

    Vincent Schram, Jean-François Tocanne & André Lopez

Authors
  1. Vincent Schram
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Jean-François Tocanne
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. André Lopez
    View author publications

    You can also search for this author in PubMed Google Scholar

Additional information

Correspondence to: A. Lopez

Rights and permissions

Reprints and permissions

About this article

Cite this article

Schram, V., Tocanne, JF. & Lopez, A. Influence of obstacles on lipid lateral diffusion: computer simulation of FRAP experiments and application to proteoliposomes and biomembranes. Eur Biophys J 23, 337–348 (1994). https://doi.org/10.1007/BF00188657

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF00188657

Key words

Search

Navigation