Controlling the aggregation and rate of release in order to improve insulin formulation: molecular dynamics study of full-length insulin amyloid oligomer models



W. M. Berhanu;A. E. Masunov


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Abbreviated Journal Title

J. Mol. Model.


Amyloid fibril; Insulin; beta-Sheet; Aggregation; Oligomer; Secondary; structure; LYQLENY; LVEALYL; Molecular dynamics simulations; Cluster; MM-GBSA; Per-residue decomposition; ALZHEIMERS A-BETA(1-40) PEPTIDE; FREE-ENERGY CALCULATIONS; FIBRIL; FORMATION; MONOMERIC INSULIN; DIABETES-MELLITUS; MASS-SPECTROMETRY; PROTEIN; MECHANISM; BINDING; SIMULATIONS; Biochemistry & Molecular Biology; Biophysics; Chemistry, ; Multidisciplinary; Computer Science, Interdisciplinary Applications


Insulin is a hormone that regulates the physiological glucose level in human blood. Insulin injections are used to treat diabetic patients. The amyloid aggregation of insulin may cause problems during the production, storage, and delivery of insulin formulations. Several modifications to the C-terminus of the B chain have been suggested in order to improve the insulin formulation. The central fragments of the A and B chains (LYQLENY and LVEALYL) have recently been identified as beta-sheet-forming regions, and their microcrystalline structures have been used to build a high-resolution amyloid fibril model of insulin. Here we report on a molecular dynamics (MD) study of single-layer oligomers of the full-length insulin which aimed to identify the structural elements that are important for amyloid stability, and to suggest single glycine mutants in the beta-sheet region that may improve the formulation. Structural stability, aggregation behavior and the thermodynamics of association were studied for the wild-type and mutant aggregates. A comparison of the oligomers of different sizes revealed that adding strands enhances the internal stability of the wild-type aggregates. We call this "dynamic cooperativity". The secondary structure content and clustering analysis of the MD trajectories show that the largest aggregates retain the fibril conformation, while the monomers and dimers lose their conformations. The degree of structural similarity between the oligomers in the simulation and the fibril conformation is proposed as a possible explanation for the experimentally observed shortening of the nucleation lag phase of insulin with oligomer seeding. Decomposing the free energy into electrostatic, van der Waals and solvation components demonstrated that electrostatic interactions contribute unfavorably to the binding, while the van der Waals and especially solvation effects are favorable for it. A per-atom decomposition allowed us to identify the residues that contribute most to the binding free energy. Residues in the beta-sheet regions of chains A and B were found to be the key residues as they provided the largest favorable contributions to single-layer association. The positive Delta Delta G(mut) values of 37.3 to 1.4 kcal mol(-1) of the mutants in the beta-sheet region indicate that they have a lower tendency to aggregate than the wild type. The information obtained by identifying the parts of insulin molecules that are crucial to aggregate formation and stability can be used to design new analogs that can better control the blood glucose level. The results of our simulation may help in the rational design of new insulin analogs with a decreased propensity for self-association, thus avoiding injection amyloidosis. They may also be used to design new fast-acting and delayed-release insulin formulations.

Journal Title

Journal of Molecular Modeling





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