The Human Islet Amyloid Polypeptide Forms Transient Membrane-Active Prefibrillar Assemblies

Abstract

The formation of amyloid fibrils by the human islet amyloid polypeptide is associated with type II diabetes. While it was previously suggested that the formed fibrils are toxic to pancreatic beta-cells due to membrane permeation activity, more recent studies suggested that protofibrillar assemblies have significantly higher potency in permeating lipid bilayers. Here, we specifically studied the membrane interaction activity of soluble and insoluble islet amyloid polypeptide assemblies at high temporal resolution. A colorimetric analysis using lipid/polydiacetylene (PDA) biomimetic vesicles clearly demonstrated the transient formation of soluble assemblies that strongly interact with the lipid vesicles. A peak in the level of membrane binding of the soluble fraction, as reflected by the colorimetric assay, was observed after incubation for approximately 1 h, followed by a decrease in the level of membrane interaction of the assemblies. The transient nature of the membrane-active assemblies was independently confirmed by a fluorescence quenching assay. Ultrastructural analysis using transmission electron microscopy provided morphological evidence of prefibrillar assemblies, supported the transient existence of membrane interacting soluble species, and facilitated observation of the non-membrane-active filaments in the solution. Taken together, our results provide experimental evidence for the formation of transient soluble prefibrillar assemblies which are highly membrane-active. The implications of these observations are discussed in light of designed fibrillization inhibitors.

Molecular Mechanisms of Amyloid Oligomers Toxicity

“…This increased conductivity was not ion specific, and thus has the potential to depolarize the membrane and lead to cellular dysfunction. A growing body of evidence points to membrane permeabilization by amyloid oligomers as a common mechanism of pathogenesis in amyloid-related degenerative diseases [13,19,[64][65][66][67][68][69][70][71][72][73][74][75][76][77]. These studies suggest that membrane permeabilization caused by amyloid oligomers is due to defects in the lipid bilayer, rather than the formation of discrete proteinaceous pores.…”

Islet amyloid polypeptide toxicity and membrane interactions

“…Pore formation has been proposed to be a key for membrane disruption (21)(22)(23), but a carpeting, detergent-like mechanism has also been advocated (24,25). There is experimental evidence that intermediates permeabilize membranes (26)(27)(28), whereas other studies implicate fibril growth at the membrane surface (29,30). The structural requirements for membrane activity are also not completely clear.…”

A hIAPP‐derived all‐d‐amino‐acid inhibits hIAPP fibrillation efficiently at membrane surface by targeting α‐helical oligomeric intermediates

“…However, compared with the study in bulk solution, the study at lipid membrane interface should be more important because hIAPP-induced toxicity in diabetes involves interactions between the cell membrane and misfolded protein. Many studies using phospholipids as model membranes have demonstrated that hIAPP binds strongly to lipid membranes [13][14][15][16][17][18][19], in particular to anionic membranes [18,20]. Moreover, the activity and mechanism of inhibition at membrane interface may be quite different from those in bulk solution.…”