Biophysical studies of membrane proteins/peptides. Interaction with ...

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signal for neighbouring cells to phagocytose the dead cell and digest it (Alberts et al., 2002). 1.4. Lipid structure and curvature Lipids in hydrated conditions exhibit polymorphic behaviour as they can assemble in different structures. The structure adopted by a lipid aggregate can be influenced by the molecular structure of the lipid itself and a myriad of environmental conditions, such as water content, pH, ionic strength, temperature and pressure. Lipid molecules can assemble in an aqueous environment either in a lamellar structure (as in a lipid bilayer) or in non-lamellar phases (micelle, hexagonal, inverted hexagonal phases, and the cubic phase (see Fig. I.4)). Because phase transitions between these different aggregates can be activated by changes in water content, these phases are called lyotropic. The different possibilities for organizations of lipids are the result of the intrinsic shape of the lipid molecule. When the headgroup of a lipid occupies the same area than the area occupied by its hydrophobic section, the leaflets or monolayers formed by association of molecules of this lipid will present zero spontaneous curvature, forming a planar membrane for which both leaflets (or monolayers) present null curvature. A lipid bilayer composed of this lipid with the same number of molecules in each monolayer should be flat. However, in order to eliminate the aqueous exposure of the hydrophobic edges of the bilayer, the bilayer can unite the edges and form a curved vesicle. In this aggregate the exterior monolayer must present a concave (or positive) curvature and the interior monolayer must present a curvature with opposite direction. In this way, spontaneous curvature of a lipid monolayer refers to the curvature observed in the absence of edge conditions (Zimmerberg, 2000). For monolayers composed of lipids presenting headgroups with a cross-section different to that of the hydrophobic tails, a spontaneous curvature will be present, and the packing of these lipids in a lipid bilayer with a lamellar structure will result in curvature stress, that can be supported by the lamellar structure only up to a certain extent. In case the lipid bilayer cannot sustain this curvature stress, the lamellar structure (L) will be broken and non-lamellar phases will arise. The basic structural phase of biological membranes is nevertheless a lamellar phospholipid bilayer matrix, and deviations from a lamellar arrangement are generally not desirable in the plasma membrane. The inclusion of lipids with propensity to non-lamellar structure leads to a 8
INTRODUCTION: BIOMEMBRANES curvature stress in the membrane, and local modulation of this stress can be achieved either by enzymatic action in the lipids, as removal of headgroups (Nieva et al., 1993) or a acyl-chain, or by protein binding, allowing local formation of non-lamellar phases. Micellar phase Hexagonal phase Lamellar phase Cubic phase Inverted hexagonal phase Figure I.4 - Representation of the most important forms of organization of lipid aggregates in an aqueous environment. Different lipid structures correspond to different lipid curvatures and are arranged in accordance to the value of the packing parameter P. Adapted from (Mouritsen, 2005) and (Seddon and Templer, 1995). The ability of a lipid to fit into a particular type of aggregate can be described by a packing parameter P, which is defined in Figure I.4. A deviation of P from 1, either positive or negative, corresponds to a deviation of the lipid shape when in the lipid aggregate from a cylindrical shape, which is the shape that fits a planar bilayer structure. Negative deviations (P < 1) correspond to hexagonal (H I ) structures and in the limit P < 1/3 to micelles. Highly phosphorylated PI can induce micelles as will be discussed in chapter IV. Positive deviations (P > 1) correspond to cubic and inverted hexagonal (H II ) aggregates. These two structures are of significant biological relevance, playing a 9
Page 1: UNIVERSIDADE TÉCNICA DE LISBOA INS
Page 4 and 5: Fernandes, F., Loura, L. M. S., Fed
Page 6 and 7: Ao Pedro e Hugo, amigos de longa da
Page 8 and 9: 2.2. - Peptides as models 2.3. - Am
Page 11 and 12: ABBREVIATIONS AND SYMBOL LIST ABBRE
Page 13: RESUMO RESUMO As biomembranas são
Page 17 and 18: SINOPSE SINOPSE Nas últimas duas d
Page 19 and 20: SINOPSE podem fornecer informação
Page 21 and 22: SINOPSE aceitantes. Dadores mais pr
Page 23 and 24: OUTLINE OUTLINE The last two decade
Page 25 and 26: OUTLINE membranes with a distributi
Page 27: OUTLINE BAR domains (tubulation of
Page 30 and 31: ion diffusion, as the energy requir
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Page 38 and 39: functional role in process such as
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Page 75 and 76: PROTEIN-PROTEIN AND PROTEIN-LIPID I
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Page 83 and 84: 2430 Biophysical Journal Volume 85
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2434 Fernandes et al. leads to an a
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2436 Fernandes et al. FIGURE 4 (A)
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2438 Fernandes et al. section of th
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2440 Fernandes et al. tein oligomer
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QUANTIFICATION OF PROTEIN-LIPID SEL
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FRET Study of Protein-Lipid Selecti
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BINDING OF INHIBITORS TO A PUTATIVE
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BINDING OF INHIBITORS TO A PUTATIVE
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INTERACTION OF THE INDOLE CLASS OF
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5272 Biochemistry, Vol. 45, No. 16,
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BINDING ASSAYS OF INHIBITORS TOWARD
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1778 F. Fernandes et al. / Biochimi
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apparently the most significant as
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110
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Introduction Quinolones are broad-s
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[9-12]. Having this into considerat
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any case, very similar, probably wi
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placement of the protein around the
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⎛ II t ⎛ R ⎞ 0 ρ () t = exp
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APPENDIX - Derivation of the FRET r
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2 2w J ( t) = '2 R − R + w / 1
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[14] L. Plançon, M. Chami, L. Lete
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acceptors) (Eqs. 4-6) (⋅-⋅-⋅)
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FIGURE 1A FIGURE 1B 130
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136
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dynamin. Soon after, the same group
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140
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142
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Introduction Control of membrane re
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Steady-state fluorescence measureme
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Rho-DOPE (acceptor). The increase o
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where η is the viscosity of the me
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ound conformation of the BAR domain
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19. Fernandes, F., L. M. S. Loura,
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Figure Legends Fig.1: N-terminal am
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FIG. 1A FIG.1B 17
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FIG. 3A FIG. 3B 19
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FIG.5A 21
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FIG 6. 23
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FIG. 8 A B 25
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The signalling functions of PI(4,5)
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172
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174
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zoxadiazol (NBD)-PC were obtained f
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Fig. 3. Fluorescence decays of diph
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CONCLUSIONS VII CONCLUSIONS Chapter
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CONCLUSIONS constants recovered by
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CONCLUSIONS Chapter IV ‐ Binding
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CONCLUSIONS Chapter VI - Clustering
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FINAL CONSIDERATIONS AND PROSPECTS
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BIBLIOGRAPHY IX BIBLIOGRAPHY Albert
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BIBLIOGRAPHY Phosphatidylinositol 4
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BIBLIOGRAPHY Glaubitz, C., Grobner,
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BIBLIOGRAPHY Koehorst, R. B. M., Sp
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BIBLIOGRAPHY Mishra, V. K., Palguna
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BIBLIOGRAPHY Pluschke, G., Hirota,
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BIBLIOGRAPHY Sperotto, M. M., and M
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BIBLIOGRAPHY Williamson, I. M., Alv
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