High‐Resolution Separation of Amyloid β‐Peptides: Structural Variants Present in Alzheimer's Disease Amyloid

: In Alzheimer's disease (AD), one of the cardinal neuropathological signs is deposition of amyloid, primarily consisting of the amyloid β‐peptide (Aβ). Structural variants of AD‐associated Aβ peptides have been difficult to purify by high‐resolution chromatographic techniques. We therefore developed a novel chromatographic protocol, enabling high‐resolution reverse‐phase liquid chromatography (RPLC) purification of Aβ variants displaying very small structural differences. By using a combination of size‐exclusion chromatography and the novel RPLC protocol, Aβ peptides extracted from AD amyloid were purified and subsequently characterized. Structural analysis by microsequencing and electrospray‐ionization mass spectrometry revealed that the RPLC system resolved a complex mixture of Aβ variants terminating at either residue 40 or 42. Aβ variants differing by as little as one amino acid residue could be purified rapidly to apparent homogeneity. The resolution of the system was further illustrated by its ability to separate structural isomers of Aβ1–40. The present chromatography system might provide further insight into the role of N‐terminally and posttranslationally modified Aβ variants, because each variant can now be studied individually.