Preparation and characterization of a highly soluble Aβ 1-42 peptide variant

Protein Expr Purif. 2019 Dec;164:105480. doi: 10.1016/j.pep.2019.105480. Epub 2019 Aug 16.

Abstract

Alzheimer's disease (AD) is a progressive neurological disease marked by the accumulation and deposition of misfolded amyloid beta or Abeta (Aβ) peptide. Two species of Aβ peptides are found in amyloid plaques, Aβ1-40 and Aβ1-42, with the latter being the more amyloidogenic of the two. Understanding how and why Aβ peptides misfold, oligomerize and form amyloid plaques requires a detailed understanding of their structure and dynamics. The poor solubility and strong aggregation tendencies of Aβ1-42 has made the isolation and characterization of its different structural isoforms (monomer, dimer, oligomer, amyloid) exceedingly difficult. Furthermore, while synthetic Aβ1-42 peptides (Aβ42syn) are readily available, the cost of isotopically labeled peptide is substantial, making their characterization by NMR spectroscopy cost prohibitive. Here we describe the design, cloning, high-level production, isotopic labeling and biophysical characterization of a modified (solubility-tagged) Aβ1-42 variant that exhibits excellent water solubility and shares similar aggregation properties as wildtype Aβ1-42. Specifically, we attached six lysines (6K) to the C-terminus of native Aβ1-42 to create a more soluble, monomeric form of Aβ1-42 called Aβ42C6K. A gene for the Aβ42C6K was designed, synthesized and cloned into Escherichia coli (E. coli) and the peptide was expressed at milligram levels. The Aβ42C6K peptide was characterized using circular dichroism (CD), NMR, electron microscopy and thioflavin T fluorescence. Its ability to form stable monomers, oligomers and fibrils under different conditions was assessed. Our results indicate that Aβ42C6K stays monomeric at high concentrations (unlike Aβ1-42) and can be induced to oligomerize and form fibrils like Aβ1-42. Our novel construct could be used to explore the structure and dynamics of Aβ1-42 as well as the interaction of ligands with Aβ1-42 via NMR.

Keywords: Alzheimer's disease; Aβ(1-42); Expression; NMR; Solubility tag.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alzheimer Disease / genetics
  • Alzheimer Disease / metabolism
  • Amyloid beta-Peptides / chemistry*
  • Amyloid beta-Peptides / genetics*
  • Amyloid beta-Peptides / metabolism
  • Amyloid beta-Peptides / ultrastructure
  • Humans
  • Mutation*
  • Peptide Fragments / chemistry*
  • Peptide Fragments / genetics*
  • Peptide Fragments / metabolism
  • Peptide Fragments / ultrastructure
  • Protein Aggregates*
  • Protein Aggregation, Pathological / genetics
  • Protein Aggregation, Pathological / metabolism
  • Protein Multimerization
  • Solubility

Substances

  • Amyloid beta-Peptides
  • Peptide Fragments
  • Protein Aggregates
  • amyloid beta-protein (1-42)

Grant support