title: Probing the nucleus model for oligomer formation during insulin amyloid fibrillogenesis.
abstract: We find evidence for a direct transition of insulin monomers into amyloid fibrils without measurable concentrations of oligomers or protofibrils, suggesting that fibrillogenesis may occur directly from assembly of denaturing insulin monomers rather than by successive transitions through protofibril nuclei. To support our finding, we obtain size distributions using electrospray differential mobility analysis (ES-DMA), which provides excellent resolution to clearly distinguish among small oligomers and rapidly generates statistically significant size distributions. The distributions detect an absence of significant peaks between 6 nm and 17 nm as the monomer reacts into fibers-exactly the size range observed by others for small-angle-neutron-scattering-measured intermediates and for circular supramolecular structures. They report concentrations in the nanomolar range, whereas our limit of detection remains three-orders-of-magnitude lower (<5 pmol/L). This finding, along with the lack of significant increases in the β-sheet content of monomers using circular dichroism, suggests monomers do not first structurally rearrange and accumulate in a β-rich state but react and reorganize at the growing fiber's tip. These results quantitatively inform reaction-based theories of amyloid fiber formation and have implications for neurodegenerative, protein conformation ailments including Alzheimer's disease and bovine spongiform encephalopathy.
authors.affiliation: National Institute of Standards and Technology, Gaithersburg, Maryland, USA. pease@eng.utah.edu
authors.forename: Leonard F
authors.initials: LF
authors.lastname: Pease
authors.forename: Mirco
authors.initials: M
authors.lastname: Sorci
authors.forename: Suvajyoti
authors.initials: S
authors.lastname: Guha
authors.forename: De-Hao
authors.initials: DH
authors.lastname: Tsai
authors.forename: Michael R
authors.initials: MR
authors.lastname: Zachariah
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Tarlov
authors.forename: Georges
authors.initials: G
authors.lastname: Belfort
mesh_terms: D000682:Amyloid / Q000737:chemistry*; D030562:Databases, Protein; D007328:Insulin / Q000737:chemistry*; D008954:Models, Biological; D008958:Models, Molecular; D020836:Protein Structure, Quaternary; D013997:Time Factors
chemical_list: D000682:Amyloid; D007328:Insulin