Quantitative Aspects of Electrochemical Detection of Amyloid‐β Aggregation

Abstract

The tyrosine based electrochemical analysis of synthetic amyloid‐β (Aβ) peptide – an analog of natural peptide implicated in Alzheimer's disease pathogenesis – was applied for a quantitative estimation of peptide aggregation in vitro. The analysis was carried out by square wave voltammetry (SWV) on carbon screen printed electrodes (SPE). The electrooxidation peak current (Ip) for Aβ42 peptide in different aggregation states was directly compared with the size and structure of Aβ42 aggregates occurring in the analyzed sample. Dynamic light scattering (DLS) and thioflavin T (ThT) based fluorescence assay were employed to estimate the size and structure of Aβ42 aggregates. The Ip was found to decrease in a linear fashion when the average diameter of aggregates and the relative ThT fluorescence in Aβ42 solutions exceeded 35 nm and 3, respectively, while being nearly constant below these values. It was suggested that the electrooxidation current is mostly generated by peptide monomers and that a depletion of the monomer pool due to inclusion of Aβ42 molecules in aggregates is responsible for the decrease of electrooxidation current. The direct electrochemistry is emerging as a method complementary to methods based on aggregates’ detection and commonly employed for monitoring Aβ aggregation. The work further enlarges the basis for application of the cost‐effective and rapid electrochemical techniques, such as SWV on carbon SPE, to in vitro studies of Aβ aggregation.

Application of an Electrochemical Method to Evaluation of Amyloid‐β Aggregation Inhibitors: Testing the RGKLVFFGR‐NH2 Peptide Antiaggregant

“…Figure demonstrates that the electrochemical oxidation of Aβ42 via the Tyr‐10 residue in phosphate buffer, measured by SWV on carbon SPE right after the addition of an equimolar amount of OR2 to the Aβ42 preparation, produces the I p at the potential practically indistinguishable from that for the Aβ42 peptide alone and lying within the range of 0.6–0.7 V ( vs . Ag/AgCl) as has been observed earlier for Aβ peptide . As expected, the OR2 peptide per se did not exhibit any electrochemical activity within the potential range tested (from 0.2 to 0.9 V) since there are no electrochemically active amino acid residues in its sequence .…”

“…As we have recently reported, I p is mostly generated by Aβ42 monomers and by relatively small (sedimentation‐resistant) Aβ42 oligomers . Thus, the decline of I p / I p (0) with time in the presence of OR2 peptide (Figure ) indicates a depletion of the pool of monomeric and oligomeric Aβ42 species, which suggests formation of large Aβ42 aggregates.…”

“…The DLS measurements were carried out on a Zetasizer Nano ZS apparatus (Malvern Instruments Ltd., UK) as described earlier . The apparatus is able to measure particle sizes in the range of 0.6 nm to 10 μm.…”

“…Aβ40 and Aβ42 oxidation peak currents ( I p ) were found to decrease with the progressive Aβ fibrillation that has been accounted for by alterations in the peptide's conformation (due to Aβ transition to the aggregated state) which affect the availability of Tyr‐10 for the oxidation on an electrode surface . Recently we have shown that the electrooxidation current of Aβ42 peptide is mostly caused by the Tyr‐10 oxidation of the ‘sedimentation‐resistant’ fraction of Aβ42 peptide molecules which includes their monomers and oligomers . The depletion of the monomers/oligomers pool due to the inclusion of Aβ42 molecules in fibrillar or amorphous aggregates is responsible for the I p decrease .…”

“…Recently we have shown that the electrooxidation current of Aβ42 peptide is mostly caused by the Tyr‐10 oxidation of the ‘sedimentation‐resistant’ fraction of Aβ42 peptide molecules which includes their monomers and oligomers . The depletion of the monomers/oligomers pool due to the inclusion of Aβ42 molecules in fibrillar or amorphous aggregates is responsible for the I p decrease .…”