Spectroscopic evidence for the existence of an obligate pre-fibrillar oligomer during glucagon fibrillation

Christensen, Peter Astrup; Pedersen, Jesper Søndergaard; Christiansen, Gunna; Otzen, Daniel Erik

doi:10.1016/j.febslet.2008.03.017

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Volume 582, Issue 9 , Pages 1341-1345, 16 April 2008

Spectroscopic evidence for the existence of an obligate pre-fibrillar oligomer during glucagon fibrillation

Edited by Jeusus Avila

Peter Astrup Christensen
- Interdisciplinary Nanoscience Center (iNANO), Centre for Insoluble Protein Structures (inSPIN), Department of Molecular Biology, University of Aarhus, Gustav Wieds Vej 10C, DK – 8000 Aarhus C, Denmark
Jesper Søndergaard Pedersen
- Interdisciplinary Nanoscience Center (iNANO), Centre for Insoluble Protein Structures (inSPIN), Department of Molecular Biology, University of Aarhus, Gustav Wieds Vej 10C, DK – 8000 Aarhus C, Denmark
Gunna Christiansen
- Institute of Medical Microbiology and Immunology, University of Aarhus, Wilhelm Meyers Allé, Bygning 1240, DK – 8000 Aarhus C, Denmark
Daniel Erik Otzen
- Interdisciplinary Nanoscience Center (iNANO), Centre for Insoluble Protein Structures (inSPIN), Department of Molecular Biology, University of Aarhus, Gustav Wieds Vej 10C, DK – 8000 Aarhus C, Denmark
- Corresponding author. Fax: +45 86 12 31 78.

Received 28 January 2008; received in revised form 6 March 2008; accepted 12 March 2008. published online 20 March 2008.

Abstract

The 29-residue peptide hormone glucagon has been used as a model system for the study of amyloid-like fibrils. Atomic force microscopy (AFM) studies have detected putative oligomeric species during this lag phase, but this has not been confirmed by any spectroscopic technique. Here we use an attached pyrene group to detect association (excimer formation) between individual glucagon molecules. Our data show that excimer formation precedes fibrillation both at different pHs and with sulfate, and support our original proposal that glucagon fibril formation is preceded by oligomer formation. We suggest that pyrene-labelling may be a useful way to monitor oligomer formation during protein fibrillation.

Keywords: Glucagon, Kinetic, Pre-fibrillar specie, Oligomer, Excimer

Abbreviations: G, glucagon, PG, N-terminal pyrene modified glucagon, ThT, thioflavin T, DLS, dynamic light scattering, AFM, atomic force microscopy, R_H, hydrodynamic radius, GdmCl, Guanidinium Chloride, FRET, Förster resonance energy transfer, TEM, transmission electron microscopy, RFU, relative fluorescence units