Early events in the fibrillation of monomeric insulin.
Insulin has a largely alpha-helical structure and exists as a mixture of hexameric, dimeric, and monomeric states in solution, depending on the conditions: the protein is monomeric in 20% acetic acid. Insulin forms amyloid-like fibrils under a variety of conditions, especially at low pH. In this study we investigated the fibrillation of monomeric human insulin by monitoring changes in CD, attenuated total reflectance-Fourier transform infrared spectroscopy, 8-anilinonaphthalenesulfonic acid fluorescence, thioflavin T fluorescence, dynamic light scattering, and H/D exchange during the initial stages of the fibrillation process to provide insight into early events involving the monomer. The results demonstrate the existence of structural changes occurring before the onset of fibril formation, which are detectable by multiple probes. The data indicate at least two major populations of oligomeric intermediates between the native monomer and fibrils. Both have significantly non-native conformations, and indicate that fibrillation occurs from a beta-rich structure significantly distinct from the native fold.
Ahmad
Atta
A
Department of Chemistry and Biochemistry, University of California, Santa Cruz, California 95064, USA.
Uversky
Vladimir N
VN
Hong
Dongpyo
D
Fink
Anthony L
AL
D019342:Acetic Acid / Q000737:chemistry; D000817:Anilino Naphthalenesulfonates / Q000494:pharmacology; D052160:Benzothiazoles; D002942:Circular Dichroism; D005456:Fluorescent Dyes / Q000494:pharmacology; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007328:Insulin / Q000737:chemistry* / Q000378:metabolism; D007700:Kinetics; D008027:Light; D013058:Mass Spectrometry; D008968:Molecular Conformation; D011487:Protein Conformation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D012542:Scattering, Radiation; D021241:Spectrometry, Mass, Electrospray Ionization; D017550:Spectroscopy, Fourier Transform Infrared; D013844:Thiazoles / Q000737:chemistry; D013997:Time Factors; D014466:Ultraviolet Rays
D000817:Anilino Naphthalenesulfonates; D052160:Benzothiazoles; D005456:Fluorescent Dyes; D007328:Insulin; D013844:Thiazoles; C009462:thioflavin T; C027132:1-anilino-8-naphthalenesulfonate; D019342:Acetic Acid