The process of amyloid-like fibril formation by methionine aminopeptidase from a hyperthermophile, Pyrococcus

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Rapid Generation of Amyloid from Native Proteins In vitro
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Rapid Generation of Amyloid from Native Proteins In vitro

Published on: December 5, 2013

K Yutani1, G Takayama, S Goda

  • 1Institute for Protein Research and Graduate School of Pharmaceutical Sciences, Osaka University, Yamadaoka, Suita, Osaka 565-0871, Japan. yutani@protein.osaka-u.ac.jp

Biochemistry
|March 8, 2000
PubMed

Insights

Methionine aminopeptidase (MAP) from P. furiosus forms amyloid-like fibrils upon denaturation. This discovery suggests even stable proteins can adopt amyloid conformations under specific denaturing conditions.

Area of Science:

  • Biochemistry
  • Protein Misfolding Diseases
  • Structural Biology

Background:

  • Amyloid fibrils are linked to severe diseases like Alzheimer's.
  • Protein misfolding and aggregation are key pathological mechanisms.
  • Understanding amyloid formation is crucial for therapeutic development.

Purpose of the Study:

  • To investigate the mechanism of amyloid fibril formation in methionine aminopeptidase (MAP) from the hyperthermophile P. furiosus.
  • To characterize the structural changes and stability of MAP during denaturation and fibril formation.

Main Methods:

  • Circular dichroism (CD) spectroscopy to monitor protein denaturation and secondary structure changes.
  • Guanidine hydrochloride (GuHCl) denaturation curves to assess protein stability.
  • Calorimetry to determine thermal stability.
  • Congo red staining and difference spectroscopy to identify amyloid characteristics.
  • X-ray diffraction and electron microscopy to analyze fibril structure and morphology.

Main Results:

  • MAP forms amyloid-like fibrils under specific denaturing conditions (3.37 M GuHCl, pH 3.31).
  • Denaturation involves loss of alpha-helical structure and gain of beta-sheet content.
  • The resulting beta-rich conformation exhibits high stability and association.
  • Amyloid-like fibrils confirmed by Congo red staining, X-ray diffraction (4.7 A reflection), and electron microscopy (1000 A length, 70 A diameter).

Conclusions:

  • Amyloid-like structures form in MAP after significant denaturation.
  • Highly stable proteins can form amyloid conformations when their denatured state is populated.
  • This finding broadens the understanding of amyloidogenesis beyond typical disease-associated proteins.

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