Rapid, Refined, and Robust Method for Expression, Purification, and Characterization of Recombinant Human Amyloid beta 1-42


Journal article


P. Prakash, Travis C. Lantz, Krupal P. Jethava, G. Chopra
Methods and Protocols, 2019

Semantic Scholar DOI PubMedCentral PubMed
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APA   Click to copy
Prakash, P., Lantz, T. C., Jethava, K. P., & Chopra, G. (2019). Rapid, Refined, and Robust Method for Expression, Purification, and Characterization of Recombinant Human Amyloid beta 1-42. Methods and Protocols.


Chicago/Turabian   Click to copy
Prakash, P., Travis C. Lantz, Krupal P. Jethava, and G. Chopra. “Rapid, Refined, and Robust Method for Expression, Purification, and Characterization of Recombinant Human Amyloid Beta 1-42.” Methods and Protocols (2019).


MLA   Click to copy
Prakash, P., et al. “Rapid, Refined, and Robust Method for Expression, Purification, and Characterization of Recombinant Human Amyloid Beta 1-42.” Methods and Protocols, 2019.


BibTeX   Click to copy

@article{p2019a,
  title = {Rapid, Refined, and Robust Method for Expression, Purification, and Characterization of Recombinant Human Amyloid beta 1-42},
  year = {2019},
  journal = {Methods and Protocols},
  author = {Prakash, P. and Lantz, Travis C. and Jethava, Krupal P. and Chopra, G.}
}

Abstract

Amyloid plaques found in the brains of Alzheimer’s disease patients primarily consists of amyloid beta 1-42 (Aβ42). Commercially, Aβ42 is synthesized using high-throughput peptide synthesizers resulting in the presence of impurities and the racemization of amino acids that affects its aggregation properties. Furthermore, the repeated purchase of even a small quantity (~1 mg) of commercial Aβ42 can be expensive for academic researchers. Here, we describe a detailed methodology for robust expression of recombinant human Aβ(M1-42) in Rosetta(DE3)pLysS and BL21(DE3)pLysS competent E. coli using standard molecular biology techniques with refined and rapid one-step analytical purification techniques. The peptide is isolated and purified from transformed cells using an optimized reverse-phase high-performance liquid chromatography (HPLC) protocol with commonly available C18 columns, yielding high amounts of peptide (~15–20 mg per 1 L culture) within a short period of time. The recombinant human Aβ(M1-42) forms characteristic aggregates similar to synthetic Aβ42 aggregates as verified by western blotting and atomic force microscopy to warrant future biological use. Our rapid, refined, and robust technique produces pure recombinant human Aβ(M1-42) that may be used to synthesize chemical probes and in several downstream in vitro and in vivo assays to facilitate Alzheimer’s disease research.


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