Fast purification of recombinant monomeric amyloid-β from E. coli and amyloid-β-mCherry aggregates from mammalian cells

Abstract

The Alzheimers disease related peptide, Amyloid-beta (A{beta})1-40 and 1-42, has proven difficult to be purified as a recombinant monomeric protein due its expression in E. coli leading to the formation of insoluble inclusion bodies and its tendency to quickly form insoluble aggregates. A vast array of methods have been used so far, yet many have pitfalls, such as the use of tags for ease of A{beta} isolation, the formation of A{beta} multimers within the time frame of extraction or the need to reconstitute A{beta} from a freeze dried state. Here, we present a rapid protocol to produce highly pure and monomeric recombinant A{beta} using a one-step ion exchange purification method and to label the peptide using a maleimide dye. The solublisation and purification steps take only three hours. We also present a protocol for the isolation of A{beta}-mCherry from mammalian cells. HighlightsO_LIPurification of untagged, monomeric recombinant A{beta} from E. coli. C_LIO_LIA fast protocol; 6 hours for E. coli growth and A{beta} expression, 2 hours to clean inclusion bodies, 45 mins to solublise and purify the peptide. C_LIO_LINo freeze drying step that can lead to oligomer formation. C_LIO_LIPurification of fluorescent A{beta}-mCherry from mammalian cells. C_LI