Materials and Methods
Peptide synthesis and preparation . Aβ40, Aβ42, single D-amino acid substituted peptides, and di-D-amino acid substituted peptides were synthesized, purified, and characterized in the Biopolymer Lab at UCLA 30 . Briefly, peptide synthesis was performed on an automated peptide synthesizer (Model 433A, Applied Biosystems, Foster City, CA) using 9-fluorenylmethoxycarbonyl-based methods. Peptides were purified using reverse phase high-performance liquid chromatography (RP-HPLC). The purity of the peptides were >95%. Quantitative amino acid analysis and mass spectrometry yielded the expected compositions and molecular weights, respectively, for each peptide. Purified peptides were stored as lyophilizates at –20°C.
Peptide lyophilizates were solvated initially in 10% (v/v) 60 mM NaOH, 45% (v/v) H2O (prepared using a Synthesis A10 water purification system (Millipore, Bedford, MA), and 45% (v/v) 22.2 mM sodium phosphate, pH 7.4, on ice. The solutions were sonicated for 1 min in an ultrasonic water bath (Model 1510, Branson Ultrasonics Corp., Danbury, CT) and then they were filtered using a Microcon centrifugal filter (30 kDa molecular mass cut-off, Millipore, Bedford, MA) at 14,000 × g for 10 min at room temperature (RT; ≈22 °C) The concentrations of the resulting filtrates were determined by UV absorbance using an extinction coefficient ε270=1276 L mol-1 cm-1. The peptide concentration then was adjusted to 40 μM using 10 mM sodium phosphate, pH 7.4.