Toward Understanding Insulin Fibrillation

Abstract

Formation of insulin fibrils is a physical process by which partially unfolded insulin molecules interact with each other to form linear aggregates. Shielding of hydrophobic domains is the main driving force for this process, but formation of intermolecular beta-sheet may further stabilize the fibrillar structure. Conformational displacement of the B-chain C-terminal with exposure of nonpolar, aliphatic core residues, including A2, A3, B11, and B15, plays a crucial role in the fibrillation process. Recent crystal analyses and molecular modeling studies have suggested that when insulin fibrillates this exposed domain interacts with a hydrophobic surface domain formed by the aliphatic residues A13, B6, B14, B17, and B18, normally buried when three insulin dimers form a hexamer. In rabbit immunization experiments, insulin fibrils did not elicit an increased immune response with respect to formation of IgG insulin antibodies when compared with native insulin. In contrast, the IgE response increased with increasing content of insulin in fibrillar form. Strategies and practical approaches to prevent insulin from forming fibrils are reviewed. Stabilization of the insulin hexameric structure and blockage of hydrophobic interfaces by addition of surfactants are the most effective means of counteracting insulin fibrillation.

Insulin analog with additional disulfide bond has increased stability and preserved activity

“…Fibril formation proceeds by partial unfolding of the insulin monomer and the C-terminus of the Bchain is thought to play an important role in this process. 37 Our results demonstrate that stabilization of the N-terminal part of the insulin molecule is sufficient to prevent fibrillation even with a flexible Bchain C-terminus, likely by stabilization of the monomeric structure, which prevents its unfolding.…”

“…37 The 4SS-insulin was tested in a Thioflavin T (ThT) assay for 45 h with vigorous shaking [ Fig. 5(B)].…”

Molecular basis for insulin fibril assembly

“…In this rare condition, full-length insulin molecules are found in fibrillar form at the site of frequent insulin injections (2)(3)(4). These insulin fibrils formed in vivo display the defining characteristics of amyloid aggregates (5), including binding the dye Congo red with ''apple-green'' birefringence, an elongated, unbranched fibrillar morphology (4), nucleation-dependent polymerization, and the cross-␤ x-ray diffraction pattern (6)(7)(8).…”

“…Recently, serum samples from patients with Parkinson's disease have been found to display an autoimmune response to insulin oligomers and fibrils (9), possibly indicating the presence of insulin aggregates in this disease as well. Insulin also forms amyloid-like fibrils in vitro, which are promoted by elevated temperatures, low pH, and increased ionic strength (4,10). In addition, insulin fibril formation has been a limiting factor in long-term storage of insulin for treatment of diabetes.…”

“…The fibrillar ␤-sheets have been described as either parallel (11)(12)(13) or antiparallel (14)(15)(16)). An early model was based on the crystal packing of a despentapeptide insulin molecule (with residues B26-B30 removed) (4,17). More recently, Jimenez et al (18) used cryo-electron microscopy images to propose that upon fibril assembly the ␣-helical insulin molecules undergo conformational transition into a flat ␤-sheet-rich state.…”