title: Early events in insulin fibrillization studied by time-lapse atomic force microscopy.
abstract: The importance of understanding the mechanism of protein aggregation into insoluble amyloid fibrils lies not only in its medical consequences, but also in its more basic properties of self-organization. The discovery that a large number of uncorrelated proteins can form, under proper conditions, structurally similar fibrils has suggested that the underlying mechanism is a general feature of polypeptide chains. In this work, we address the early events preceding amyloid fibril formation in solutions of zinc-free human insulin incubated at low pH and high temperature. Here, we show by time-lapse atomic force microscopy that a steady-state distribution of protein oligomers with a quasiexponential tail is reached within a few minutes after heating. This metastable phase lasts for a few hours, until fibrillar aggregates are observable. Although for such complex systems different aggregation mechanisms can occur simultaneously, our results indicate that the prefibrillar phase is mainly controlled by a simple coagulation-evaporation kinetic mechanism, in which concentration acts as a critical parameter. These experimental facts, along with the kinetic model used, suggest a critical role for thermal concentration fluctuations in the process of fibril nucleation.
authors.affiliation: Istituto Nazionale per la Fisica della Materia, Dipartimento di Fisica, and Cimaina, Università di Milano, Milan, Italy.
authors.forename: Alessandro
authors.initials: A
authors.lastname: Podestà
authors.forename: Guido
authors.initials: G
authors.lastname: Tiana
authors.forename: Paolo
authors.initials: P
authors.lastname: Milani
authors.forename: Mauro
authors.initials: M
authors.lastname: Manno
mesh_terms: D000682:Amyloid / Q000737:chemistry; D001703:Biophysics / Q000379:methods*; D003461:Crystallography; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007328:Insulin / Q000737:chemistry* / Q000378:metabolism; D007700:Kinetics; D008027:Light; D018625:Microscopy, Atomic Force / Q000379:methods*; D010455:Peptides / Q000737:chemistry; D012542:Scattering, Radiation; D013696:Temperature; D013816:Thermodynamics; D013997:Time Factors; D015032:Zinc / Q000737:chemistry
chemical_list: D000682:Amyloid; D007328:Insulin; D010455:Peptides; D015032:Zinc