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

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sphingomyelin, the most abundant sphingolipid, which is a significant component of animal plasma membranes. 1.2.1.3. Glycolipids Both glycerolipids and sphingolipids can be linked to a carbohydrate in their headgroups, giving rise to a glycolipid. Glycolipids play important roles in the interactions of the cell with its surroundings. Glycoglycerolipids are very abundant in chloroplast membranes, as well as in blue algae and bacteria. They are however rarely found in animals (Gennis, 1989). In glycosphingolipids, one or more carbohydrates are linked to the terminal hydroxyl group of the sphingosine backbone. The simplest form of glycosphingolipid is a cerebroside that contains a single carbohydrate residue, either glucose or galactose. Complex glycosphingolipids called gangliosides present one or two branched carbohydrate chains containing sialic acid groups. Glycosphingolipids are as a rule located in the outer surface of the plasma membrane with varying concentrations (2-10 %), and are particularly abundant in the nervous system. 1.2.1.4. Sterols The basic structure of sterols is a four ring hydrocarbon. This structure confers the molecule much more rigidity than other lipids. The most important sterol in animal membranes is cholesterol (Fig. I.3). In cholesterol, a hydrocarbon chain is linked to one end of the ring system and a hydroxyl group is attached to the other end of the structure, this being the polar headgroup of the molecule. In the lipid bilayer the molecule is oriented parallel to the fatty-acid chains of other lipids while the hydroxyl group interacts with nearby headgroups. Cholesterol is only found in eukaryotes, particularly in animal plasma membranes, lysosomes, endosomes and Golgi apparatus. The concentration of cholesterol is especially high in the animal plasma membrane (20 - 30 %). In plants cholesterol exists in low amounts, and other sterols are present, namely sitoesterol and stigmasterol. Ergosterol is found in yeast and other eukaryotic microorganisms. 6
INTRODUCTION: BIOMEMBRANES Figure 1.3 – Structure of cholesterol (From Berg et al., 2002). 1.3. Lipid asymmetry across the bilayer Lipid composition in the cytoplasmatic and exoplasmatic leaflets of the bilayer is asymmetric. In plasma membranes, the cytoplasmatic leaflet is enriched in PE, PS, and PI, while in the exoplasmatic leaflet lipids like PC, glycolipids, sphingomyelin and cholesterol exist in higher concentrations. One of the factors contributing to this lipid asymmetry is the site of synthesis of the lipids in the membranes of the endoplasmic reticulum and Golgi, as it dictates the plasma membrane leaflet where the lipid is to be inserted. In this way, addition of carbohydrates to glycolipids is done through the luminal side of the Golgi apparatus that is topologically equivalent to the exterior of the cell. This however does not account for all asymmetry observed and the action of ATP-powered transport proteins called translocases is essential (Lodish et al., 2000). However, asymmetry would be lost if lipids easily crossed from one leaflet to the other. In fact, in protein free liposomes, the kinetics of this process is extremely slow, on the order of hours/days. This is due to the extremely energetically unfavourable process of inserting the polar headgroup of a lipid in the hydrophobic membrane interior. Lipid asymmetry is of functional importance, and many cytosolic proteins bind to specific lipid headgroups (like PS or PI) in the cytosolic leaflet of the bilayer. Animals also use phospholipid asymmetry in the plasma membrane of cells as a control to distinguish between dead and healthy cells, as when animal cells undergo programmed cell death, or apoptosis. In this case, PS lipids that are normally found enriched in the cytosolic leaflet, distribute between both leaflets of the bilayer. This functions as a 7
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
Page 32 and 33: acid. If no more groups are linked
Page 36 and 37: signal for neighbouring cells to ph
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Page 58 and 59: changes abruptly in the interfacial
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Page 62 and 63: some α-helical membrane proteins a
Page 64 and 65: The formation of a lipid population
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Page 75 and 76: PROTEIN-PROTEIN AND PROTEIN-LIPID I
Page 77 and 78: PROTEIN-PROTEIN AND PROTEIN-LIPID I
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Page 83 and 84: 2430 Biophysical Journal Volume 85
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2432 Fernandes et al. Coat protein
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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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