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Conformational intermediate of the amyloidogenic protein beta 2-microglobulin at neutral pH
N H Heegaard1, J W Sen, N C Kaarsholm
1Department of Autoimmunology, Statens Serum Institut, 2300 Copenhagen S, Denmark. nhe@ssi.dk
Abstract:
Aggregation and fibrillation of beta(2)-microglobulin are hallmarks of dialysis-related amyloidosis. We characterize perturbations of the native conformation of beta(2)-microglobulin that may precede fibril formation. For a beta(2)-microglobulin variant cleaved at lysine 58, we show using capillary electrophoresis that two conformers spontaneously exist in aqueous buffers at neutral pH. Upon treatment of wild-type beta(2)-microglobulin with acetonitrile or trifluoroethanol, two conformations were also observed. These conformations were in equilibrium dependent on the sample temperature and the percentage of organic solvent present. Circular dichroism showed a loss of beta-structures and gain of alpha-helices. Reversal to the native conformation occurred when removing the organics. Affinity capillary electrophoresis experiments showed increased specific interactions of the nonnative beta(2)-microglobulin conformation with the dyes 8-anilino-1-naphthalene sulfonic acid and Congo red. The observations may relate to early folding events prior to amyloid fibrillation and facilitate the development of methods to detect and inhibit pro-amyloid protein and peptide conformations.
Insights
Dialysis-related amyloidosis involves beta(2)-microglobulin fibrillation. This study identifies nonnative protein conformations that may precede amyloid formation, offering insights for detection and inhibition strategies.
Area of Science:
- Biochemistry
- Protein Misfolding Diseases
- Structural Biology
Background:
- Dialysis-related amyloidosis is characterized by beta(2)-microglobulin (β2M) aggregation and fibrillation.
- Understanding the early conformational changes in β2M is crucial for elucidating amyloid formation pathways.
Purpose of the Study:
- To characterize nonnative conformations of beta(2)-microglobulin (β2M) that may precede amyloid fibril formation.
- To investigate the conditions and properties of these transient protein states.
Main Methods:
- Capillary electrophoresis to detect and separate protein conformers.
- Circular dichroism spectroscopy to analyze secondary structure changes.
- Affinity capillary electrophoresis to assess interactions with amyloid-binding dyes.
Main Results:
- Two distinct conformers of β2M were observed in aqueous buffers and upon treatment with organic solvents.
- These nonnative conformations exhibited altered secondary structures (loss of β-sheets, gain of α-helices).
- Nonnative β2M showed increased specific interactions with dyes like Congo red, indicative of amyloidogenic properties.
Conclusions:
- Pre-fibrillar, nonnative conformations of β2M exist and can be induced by environmental factors.
- These findings provide insights into early events in amyloidogenesis and potential targets for therapeutic intervention.
- The study facilitates the development of methods to detect and inhibit pro-amyloidogenic protein conformations.