A λ-Dynamics Investigation of Insulin Wakayama and Other A3 Variant Binding Affinities to the Insulin Receptor.

Abstract

Insulin Wakayama is a clinical insulin variant where a conserved valine at the third residue on insulin's A chain (Val^A3) is replaced with a leucine (Leu^A3), weakening insulin receptor (IR) binding by 140-500-fold. This severe impact on binding from a subtle modification has posed an intriguing problem for decades. Although experimental investigations of natural and unnatural A3 mutations have highlighted the sensitivity of insulin-IR binding at this site, atomistic explanations of these binding trends have remained elusive. We investigate this problem computationally using λ-dynamics free energy calculations to model structural changes in response to perturbations of the Val^A3 side chain and to calculate associated relative changes in binding free energy (ΔΔG_bind). The Wakayama Leu^A3 mutation and seven other A3 substitutions were studied in this work. The calculated ΔΔG_bind results showed high agreement compared to experimental binding potencies with a Pearson correlation of 0.88 and a mean unsigned error of 0.68 kcal/mol. Extensive structural analyses of λ-dynamics trajectories revealed that critical interactions were disrupted between insulin and the insulin receptor as a result of the A3 mutations. This investigation also quantifies the effect that adding an A3 C_δ atom or losing an A3 C_γ atom has on insulin's binding affinity to the IR. Thus, λ-dynamics was able to successfully model the effects of mutations to insulin's A3 side chain on its protein-protein interactions with the IR and shed new light on a decades-old mystery: the exquisite sensitivity of hormone-receptor binding to a subtle modification of an invariant insulin residue.