Alzheimer's disease (AD) is characterized by the aggregation and extracellular deposition of the Aβ(1-40) and Aβ(1-42) peptides as amyloid plaques. Numerous studies suggest that the Aβ pathologic effects are related to the conversion of monomeric, random structured Aβ into aggregated β-sheet structures that eventually form plaques. This thesis focuses on two projects that are involved with inhibition of the Aβ aggregation and plaque formation processes.
The first project compared the methionine-35 (Met35)oxidation rates of the Aβ(1-40) and Aβ(1-42) peptides. Previous work demonstrated that oxidation of the Met35 side-chain to the sulfoxide (Met35red to Met35ox) impedes aggregation and formation of amyloid protofibrils. We and others hypothesized that the monomeric Aβ peptide may serve a beneficial role and normally function as an antioxidant. NMR and absorbance studies demonstrated that the Aβ(1-42) is more sensitive to oxidation with hydrogen peroxide (a natural brain oxidant) than the Aβ(1-40). However, because the oxidation rates of both peptides were slow relative to other biological substrates, it may happen that the Aβ peptides do not function as antioxidants under normal physiologic conditions. During the course of the experiments, we discovered that HPLC purification of the Aβ peptides under standard acidic conditions (water, acetonitrile, trifluoroacetic acid) promoted the Met35red to Met35ox conversion, so a modified basic solvent system (water, acetonitrile, ammonium hydroxide) was developed.
The second project studied antibodies raised against the potato virus-Y that bind to the Aβ peptide. Fluorescence spectroscopy demonstrated that the binding constant was
in the micromolar range, plus NMR and circular dichroism revealed that the antibody induced only minor changes in the Aβ peptide secondary structure. The antibody prevented NMR signal loss (a normal consequence associated with Aβ aging and aggregation), and Thioflavin T binding experiments revealed that sub-molar ratios of the antibody inhibited aggregation. Together, these data suggest that, although weak, the potato virus-Y antibody binding prevents Aβ aggregation, and may constitute a starting point for the design of new immunotherapeutic ligands for treatment of AD.