The Changing Face of Glucagon Fibrillation: Structural Polymorphism and Conformational Imprinting

We have established a time-resolved fluorescence assay to study fibrillation of the 29 residue peptide hormone glucagon under a variety of different conditions in a high-throughput format. Fibrils formed at pH 2.5 differ in fibrillation kinetics, morphology, thioflavin T staining and FTIR/CD spectra...

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Bibliographic Details
Published in:Journal of molecular biology 2006-01, Vol.355 (3), p.501-523
Main Authors: Pedersen, Jesper Søndergaard, Dikov, Dantcho, Flink, James L., Hjuler, Hans Aage, Christiansen, Gunna, Otzen, Daniel Erik
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Biopolymers - chemistry
Circular Dichroism
Dimerization
fibrillation
Glucagon - chemistry
glucagons
Hydrogen-Ion Concentration
imprinting
Kinetics
Models, Biological
Molecular Sequence Data
polymorphism
Protein Conformation
Sodium Chloride - chemistry
Spectroscopy, Fourier Transform Infrared
Temperature
Urea - chemistry
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Summary:We have established a time-resolved fluorescence assay to study fibrillation of the 29 residue peptide hormone glucagon under a variety of different conditions in a high-throughput format. Fibrils formed at pH 2.5 differ in fibrillation kinetics, morphology, thioflavin T staining and FTIR/CD spectra depending on salts, glucagon concentration and fibrillation temperature. Apparent fibrillar stability correlates with spectral and kinetic properties; generally, fibrils formed under conditions favourable for rapid fibrillation (ambient temperatures, high glucagon concentration or high salt concentration) appear less thermostable than those formed under more challenging conditions (high temperatures, low glucagon or low salt concentrations). Properties of preformed fibrils used for seeding are inherited in a prion-like manner. Thus, we conclude that the structure of fibrils formed by glucagon is not the result of the global energy minimization, but rather kinetically controlled by solvent conditions and seed-imprinting. Fibrillar polymorphism, which is being reported for an increasing number of proteins, probably reflects that fibrils have not been under evolutionary constraints to retain a single active conformation. Our results highlight the complexity of the fibrillation mechanism of glucagon, since even subtle changes in fibrillation conditions can alter the type of fibrils formed, or result in formation of mixtures of several types of fibrils.
ISSN:0022-2836
1089-8638
DOI:10.1016/j.jmb.2005.09.100