A Biotechnological Method Provides Access to Aggregation Competent Monomeric Alzheimer's 1–42 Residue Amyloid Peptide

Abstract

Senile plaques, a neuropathological hallmark of Alzheimer's disease, consist primarily of insoluble aggregates of ß-amyloid peptide (Aß). A 42-residue peptide (Aß_1-42) appears to be the predominant form. In contrast to Aß_1-40, Aß_1-42 is characterized by its extreme tendency to aggregate into fibers or precipitate. A tailored biotechnological method prevents aggregation of Aß_1-42 monomers during its production. The method is based on a protein tail fused to the amino terminus of Aß. This tail leads to a high expression in E. coli, and a histidine affinity tag facilitates purification. Selective cleavage of the fusion tail is performed with cyanogen bromide by immobilizing the fusion protein on a reversed phase chromatography column. Cleavage then occurs only at the methionine positioned at the designed site but not at the methionine contained in the membrane anchor sequence of Aß. Furthermore, immobilization prevents aggregation of cleaved Aß. Elution from the HPLC column and all succeeding purification steps are optimized to preserve Aß_1-42 as a monomer. Solutions of monomeric Aß_1-42 spontaneously aggregate into fibers within hours. This permits the investigation of the transition of monomers into fibers and the correlation of physico-chemical properties with biological activities. Mutations of Aß_1-42 at postion 35 influence the aggregation properties. Wild-type Aß_1-42 with methionine at position 35 has similar properties as Aß with a methionine sulfoxide residue. The fiber formation tendency, however, is reduced when position 35 is occupied by a glutamine, serine, leucine, or a glutamic acid residue.