SPECTROSCOPY, IMAGING, OTHER TECHNIQUES |
Early Stages of Amyloid Fibril Formation Studied by Liquid-State NMR: The Peptide Hormone Glucagon
Anna Sigrid Pii Svane 1, Kasper Jahn 2, Taru Deva 2, Anders Malmendal 2, Daniel Erik Otzen 2, Jens Dittmer 2 and Niels Chr. Nielsen 2*
1 University of Aarhyus
2 University of Aarhus
* To whom correspondence should be addressed. E-mail: ncn{at}chem.au.dk.
Submitted on September 27, 2007
Revised on October 24, 2007
Accepted on 21 February 2008
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Abstract |
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The 29-residue peptide hormone glucagon forms amyloid fibrils within a few hours at low pH. In this study, we use glucagon as a model system to investigate fibril formation by liquid-state 1H NMR spectroscopy One-dimensional as well as correlation and diffusion experiments monitoring the fibril formation process provide insight into the early stages of the pathway, on which the molecules aggregate to fibrils. In conjunction with this, exchange experiments give information about the end-state conformation. Within the limits of detection there are no signs of larger oligomeric intermediates in the course of the fibril formation process. Kinetic information is extracted from the time course of the signal decay of the residual free glucagons. This suggests that glucagon amyloids form by a nucleated growth mechanism, in which trimers (rather than monomers) of glucagon interact directly with the growing fibrils, as opposed to interacting with each other. The results of proton/deuterium exchange experiments on mature fibrils with subsequent dissolution show that the N-terminal of glucagon is least amenable to exchange, which indicates that this part is strongly involved in the intermolecular bonds of the fibrils.
Key Words:
AFM, Fibril formation, Glucagon, liquid-state NMR
