A tale of two tails: Self-assembling properties of A- and B-chain parts of insulin's highly amyloidogenic H-fragment.

Due to the spontaneous transition of native insulin into therapeutically inactive amyloid, prolonged storage decreases effectiveness of the hormone in treatment of diabetes. Various regions of the amino acid sequence have been implicated in insulin aggregation. Here, we focus on smaller fragments of the highly amyloidogenic H-peptide comprising disulfide-bonded N-terminal sections of insulin's A-chain (13 residues) and B-chain (11 residues). Aggregation patterns of N-terminal fragments of A-chain (ACC1-13, ACC1-11, ACC6-13, ACC6-11, all retaining Cys6A-Cys11A disulfide bond) and B-chain (B1-11(7A)) are examined at acidic and neutral pH. ACC1-11 is the smallest fragment found to be amyloidogenic at either pH; removal of the N-terminal GIVEQ section renders this fragment entirely non-amyloidogenic. The self-assembling properties of ACC1-11 contrast with aggregation-resistant behavior of B1-11(7A) and its disulfide-linked homodimer, (B1-11)2 aggregating only at neutral pH. Fibrillar ACC1-11 is similar to insulin amyloid in terms of morphology and infrared features. Secondary nucleation is likely to account for the detected shortening of insulin aggregation lag phase at neutral pH upon cross-seeding with pre-formed fibrils of ACC1-11 or (B1-11)2. An aggregation-enhancing effect of monomeric ACC1-11 on co-dissolved native insulin is also observed. Our findings are discussed in the context of mechanisms of insulin aggregation.