| Increasing evidence indicates that soluble aggregates of amyloid beta protein (Abeta) are neurotoxic and their interaction with membranes is vital to Alzheimer's disease (AD). However, studies of Abeta aggregation in solution and its interaction with membranes are impeded by difficulty in isolating these unstable, dynamic oligomers. On the other hand, though certain osmolytes were shown to delay aggregation in vitro and reduce Abeta neurotoxicity, their effects on aggregation versus membrane interactions have not been well studied, and their mechanisms are poorly understood.;In this dissertation, hydrogen-deuterium exchange (HX) detected by mass spectrometry (MS) was used to measure Abeta(1-40) aggregate distributions without pretreatment that might alter the aggregates structure or distribution. Different peaks in the mass spectra were assigned to monomer, low molecular weight oligomer (LMW), high molecular weight oligomer (HMW), and fibril based on distinct HX labeling behaviors, with complementary circular dichroism (CD) and Congo red binding data. HX-MS also showed that the HMW dissociated much more slowly than LMW in fresh sample. The amount of HMW deconvoluted from mass spectra was well correlated with Abeta toxicity previously reported. The interaction between Abeta aggregates and membrane lipids was studied via dye leakage assay, HX-MS, CD and fluorescence spectroscopy. Severe dye leakage was observed for oligomer abundant samples. At the same time, the lipid-peptide interaction accelerated transition from HMW to fibril was also detected. Fluorescence quenching of AMCA-Abeta samples revealed protection of the N-terminally positioned fluoroprobe when liposomes were added. Taken together, the evidence supports the conclusion that the interaction between liposome and Abeta oligomers increases membrane permeability and accelerates Abeta fibrillogenesis.;The influence of three osmolytes on Abeta aggregation and Abeta-membrane interactions were also studied. For Abeta aggregation in absence of liposomes, trehalose and sucrose delayed Abeta aggregation substantially, but not trimethylamine N-oxide (TMAO). The markedly different influences of disaccharides versus TMAO might be attributed to their varied affinity with peptide backbone, which is consistent with the trend of noncovalent osmolyte-Abeta complex adduct formation during the electrospray ionization process. In contrast and surprisingly, all three osmolytes significantly attenuated dye leakage, as well as inhibited the lipid-accelerated Abeta aggregation. |
