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Blocking PSD95-PDZ3's amyloidogenesis through point mutations that inhibit high-temperature reversible
Tomonori Saotome1,2,3, Sawaros Onchaiya1, Subbaian Brindha1
1Department of Biotechnology and Life Science, Tokyo University of Agriculture and Technology, Japan.
The FEBS Journal
|December 30, 2021
Summary
High temperatures trigger amyloid formation in postsynaptic density protein 95 PDZ3. Suppressing reversible oligomerization significantly inhibits this amyloidogenesis, revealing a key role for oligomers in fibril formation.
Area of Science:
- Biochemistry
- Structural Biology
- Protein Misfolding
Background:
- The third PDZ domain of postsynaptic density protein 95 (PSD95-PDZ3) is a small globular protein.
- PSD95-PDZ3 exhibits unusual three-state thermal unfolding with reversible oligomerization (RO) at high temperatures.
- This high-temperature RO is uncharacteristic for small globular proteins and linked to amyloid fibril formation.
Purpose of the Study:
- To investigate the role of high-temperature reversible oligomerization (RO) in the amyloidogenesis of PSD95-PDZ3.
- To determine if suppressing RO affects amyloid formation.
- To understand the relationship between protein unfolding, oligomerization, and amyloidogenicity.
Main Methods:
- Protein engineering: Created variants (F340A, L342A) to suppress RO and single-alanine mutants.
- Structural and oligomerization analysis: Circular Dichroism (CD), Analytical Ultracentrifuge (AUC), Differential Scanning Calorimetry (DSC).
- Amyloidogenicity assessment: Thioflavin T (ThT) fluorescence and Transmission Electron Microscopy (TEM).
Main Results:
- Mutations suppressing high-temperature RO strongly inhibited PSD95-PDZ3 amyloidogenesis.
- Variants that formed RO under high-temperature conditions also formed amyloid fibrils.
- Native structure was retained at ambient temperature for all variants.
Conclusions:
- High-temperature reversible oligomerization is a critical intermediate step in PSD95-PDZ3 amyloid formation.
- Targeting or preventing RO could be a strategy to inhibit amyloidogenesis in this protein.
- The findings provide insights into the mechanism of amyloid formation in globular proteins.
