Membrane Destabilization by Alzheimer's Amyloid β Peptide

Alzheimer's Amyloid-β peptide (Aβ) is a 39-42 amino acid fragment of the amyloid precursor protein. Accumulation of the neurotoxic Aβ in the brains of Alzheimer's patients forms detrimental plaques. Another mechanism of neuronal damage is membrane destabilization by insertion of the peptide into brain cell membranes and disruption of ionic homeostasis. The molecular mechanism of membrane permeabilization by Aβ remains largely uncharacterized. Experiments with live cells and lipid membranes have identified ion channels formed by Aβ. The channel formed by an 11 amino acid residue fragment, Aβ25-35, has been modeled by Molecular Dynamics simulations as an 8-stranded β-barrel, but this structure has not been supported by direct experimental data. The objective of this work was to study the mechanism of membrane permeabilization by Aβ using biophysical techniques. Unilamellar lipid vesicles were prepared in a buffer containing 30 mM CaCl2, followed by removal of calcium from the external medium via a desalting column and addition of 0.1 mM Quin-2, a fluorescent calcium indicator. Before peptide addition, the vesicle sample showed little fluorescence because of spatial sequestration of calcium and Quin-2. Addition of submicromolar concentration of the Aβ25-35 peptide resulted in gradual increase in Quin-2 fluorescence, indicating formation of Ca2+-permeable pores in vesicle membranes. Addition of higher concentrations of the peptide resulted in instantaneous increase in fluorescence, comparable to the effect of detergents like Triton X-100, indicating rupture of vesicle membranes. Thus, the data suggest membrane pore formation at low peptide concentrations and membrane disruption at higher concentrations. These data are consistent with a mechanism of detergent-like action of the Aβ peptide; at concentrations below the aggregation threshold (similar to critical micelle concentration) it forms membrane pores, and at higher concentration it acts like a detergent and solubilizes the membranes of surrounding cells.