The interaction of amyloid-β peptide with lipid membranes and glycosaminoglycans

The aim of the thesis is a better understanding of the thermodynamics and structural aspects of the interaction of Aβ(1-40) with lipid membranes and glycosaminoglycans (GAGs). We investigate the thermodynamic driving forces of the random-coil-to-β-structure transition of Aβ(1-40) in a membrane environment. In contrast to previous reports, we use anionic lipid vesicles coated with the polymer poly(ethylenglycol) (PEG), which prevents vesicle aggregation. Circular dichroism (CD) spectroscopy demonstrate that PEGylated vesicles induce a random-coil-to-β-structure transition with a defined conformational endpoint in Aβ(1-40) and four double d-isomers. The additional transition to α-helix of Aβ(1-40), reported in earlier studies for high lipid-to-peptide molar ratios of non-PEGylated anionic vesicles, was diminished. Hence, the defined conformational endpoint allows for the quantification of the membrane-induced β-structure, which is dependent on the location of the double-d substitution in the peptide sequence. Isothermal titration calorimetry (ITC) reveals the thermodynamic binding parameters for the studied peptides. The thermodynamic parameters of the β-structure formation are deduced by correlating the structural data with the thermodynamic binding parameters. In addition, we study the effect of pH on the thermodynamic binding parameters of Aβ(1-40).
In order to elucidate the role of GAGs in the Aβ(1-40) fibrillization process, we investigate the interaction of Aβ(1-40) with heparin. First, CD measurements demonstrate the effect of physico-chemical parameters, including pH, ionic strength and temperature, on the conformational state of Aβ(1-40). These measurements lead to experimental conditions, in which the interaction of monomeric Aβ(1-40) with heparin can be studied. The monomeric state of Aβ(1-40) is a prerequisite for a full thermodynamic characterization of the interaction by ITC. Under the given conditions, ITC experiments demonstrate that Aβ(1-40) interacts
with high affinity with heparin. CD spectroscopy reveals a structural transition of Aβ(1-40) from random-coil to β-structure. By studying the double-d isomers of Aβ(1-40), we identify a molecular pattern of the Aβ(1-40)-heparin interaction. We predict the binding constant for the Aβ(1-40)-heparin interaction at physiological ionic strength by using the oligolysine model. Finally, CD measurements reveal that GAGs do not induce a structural transition of Aβ(1-40) at physiological pH.
In the last part of the thesis, the interaction of Aβ(1-40) to cationic lipid vesicles is studied. CD spectroscopy measurements demonstrate that Aβ(1-40) undergoes two sequential structural transitions upon the binding to cationic membranes. First, a random-coil to β-structure transition is observed, followed by a transition to an α-helix at high lipid-to-peptide molar ratios. Compared to anionic lipid vesicles, the membrane-induced β-structure of Aβ(1-40) is less pronounced for cationic membranes, mainly due to the higher pH value at the membrane surface. The thermodynamics of the Aβ(1-40) membrane binding is studied with ITC for three buffer conditions, namely Tris-HCl, Mops and Hepes. Dynamic light scattering measurements show that the binding of Aβ(1-40) induces the formation of large