Expression and purification of a recombinant amyloidogenic peptide from transthyretin for solid-state NMR spectroscopy

Philippe S. Nadaud, Mohosin Sarkar, Bo Wu, Cait E. MacPhee, Thomas J. Magliery, Christopher P. Jaroniec

Research output: Contribution to journalArticlepeer-review

Abstract / Description of output

We describe the expression and purification of a model amyloidogenic peptide comprising residues 105-115 of human transthyretin (TTR105-115). Recombinant TTR105-115, which does not contain any non-native residues, was prepared as part of a fusion protein construct with a highly soluble B1 immunoglobulin binding domain of protein G (GB1), with typical yields of similar to 4 mg/L of uniformly C-13,N-15-enriched HPLC-purified peptide per liter of minimal media culture. Amyloid fibrils formed by recombinant TTR105-115 were characterized by transmission electron microscopy and solid-state NMR spectroscopy, and found to be comparable to synthetic TTR105-115 fibrils. These results establish recombinant TTR105-115 as a valuable model system for the development of new solid-state NMR techniques for the atomic-level characterization of amyloid architecture. (C) 2009 Elsevier Inc. All rights reserved.

Original languageEnglish
Pages (from-to)101-108
Number of pages8
JournalProtein Expression and Purification
Volume70
Issue number1
DOIs
Publication statusPublished - Mar 2010

Keywords / Materials (for Non-textual outputs)

  • Fusion protein
  • GB1
  • Recombinant human transthyretin 105-115
  • Escherichia coli BL21(DE3)
  • Ni2+ affinity chromatography
  • Amyloid fibrils
  • Solid-state NMR spectroscopy
  • ALPHA-SYNUCLEIN FIBRILS
  • BETA-SHEET STRUCTURE
  • HUMAN PRION PROTEIN
  • ANGLE-SPINNING NMR
  • ESCHERICHIA-COLI
  • HIGH-RESOLUTION
  • MOLECULAR-CONFORMATION
  • STRUCTURAL MODEL
  • RESIDUES
  • SYSTEM

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