An improved method of preparing the amyloid beta-protein for fibrillogenesis and neurotoxicity experiments

Synthetic amyloid beta-protein (A beta) is used widely to study fibril form ation and the physiologic effects of low molecular weight and fibrillar for ms of the peptide on cells in culture or in experimental animals. Not infre quently, conflicting results have arisen in these studies, in part due to v ariation in the starting conformation and assembly state of A beta. To avoi d these problems, we sought a simple, reliable means of preparing A beta fo r experimental use. We found that solvation of synthetic peptide with sodiu m hydroxide (A beta-NaOH) followed by lyophilization, produced stocks with superior solubility and fibrillogenesis characteristics. Solubilization of the pretreated material with neutral buffers resulted in a pH transition fr om similar to 10.5 to neutral, avoiding the isoelectric point of A beta (pI approximate to 5.5), at which A beta precipitation and aggregation propens ity are maximal. Relative to trifluoroacetate (A beta-TFA) or hydrochloric acid (A beta-HCl) salts of A beta, yields of "low molecular weight A beta" (monomers and/or dimers) were improved significantly by NaOH pretreatment. Time-dependent changes in circular dichroism spectra and Congo red dye-bind ing showed that A beta-NaOH formed fibrils more readily than did the other A beta preparations and that these fibrils were equally neurotoxic. NaOH pr etreatment thus offers advantages for the preparation of A beta for biophys ical and physiologic studies.