Expression, purification, and characterization of recombinant human β-amyloid42 peptide in Escherichia coli

Alzheimer’s disease (AD) is a neurodegenerative disorder characterized by a progressive loss of cognitive function. Evidence indicates that abnormal processing and extracellular deposition of the β-amyloid42 peptide, the longer form of proteolytic derivative of the transmembrane glycoprotein–amyloid precursor protein (APP), is a key step in the pathogenesis of AD. Since it is convenient and economical to obtain such a peptide biologically, in this study, we report for the first time a method to express in E. coli and purify β-amyloid42 using glutathione-S-transferase (GST) fusion system. β-Amyloid42 gene was inserted into a vector pGEX-4T-1 to construct a GST-fusion protein. The fusion protein GST-β-amyloid42, expressed in BL21 (DE3) strain, was purified with GSH-affinity chromatography followed by thrombin cleavage. The digested product was further purified with an additional GSH-affinity and a Benzamidine chromatography step. After cleavage and purification, the β-amyloid42 moiety showed the expected size of 4.5 kDa on Tricine–SDS–PAGE, and was further confirmed by Western blot. Moreover, the fibrillar recombinant β-amyloid42 exhibited great aggregation activity and showed neurotoxicity on neuron cells in vitro. These results suggest that our method will be useful in obtaining a large quantity of recombinant β-amyloid42 peptide for further physiological and biochemical studies.