Unveiling molecular mechanism underlying inhibition of human islet amyloid polypeptide fibrillation by benzene carboxylic acid-peptide conjugate

Abstract

Type 2 diabetes (T2D) is linked to the apoptosis of insulin-producing β-cells due to the aberrant fibrillation of the human islet amyloid polypeptide (hIAPP or amylin) to cytotoxic aggregates. Profit et al. generated conjugates (C1–C7) by appending benzene carboxylic acids of varying charges to the N-terminal of hIAPP_22-29 fragment peptide NFGAILSS to modulate hIAPP fibrillation and cytotoxicity. C5 (4 µM) derived by conjugating low-cost, commercially available benzene-1,2,4,5-tetracarboxylic acid known as pyromellitic acid to NFGAILSS completely abolishes the hIAPP (40 µM) self-assembly as noted in the thioflavin T (ThT) fluorescence assay. The circular dichroism (CD) spectra highlighted that C5 stabilized hIAPP in a distinctive conformation and blocked its conformational switching to amyloidogenic β-sheet structure. C5 possessing a charge-dense pyromellitic acid moiety appended on the N-terminal region of self-recognition hIAPP fragment sequence NFGAILSS created significant interest as it effectively inhibited hIAPP fibrillation and possessed lower molecular mass, smaller size, and charge as compared to hIAPP aggregation inhibitor EEEENFGAILSS (P10). However, it remains unclear how C5 traps hIAPP into a unique conformation that abolishes its self-aggregation propensity. Thus, molecular dynamics (MD) simulations have been employed to illuminate the conformational transitions and structural changes in hIAPP on the inclusion of C5. C5 displayed high-affinity binding interactions to hIAPP (ΔG_binding = −37.11 ± 3.72 kcal/mol) with major contributions from the van der Waals and electrostatic interactions. Furthermore, residue-specific binding free energy analysis depicted high-affinity binding interactions of C5 with Phe15, His18, Asn21, and Tyr37 of hIAPP that play a crucial role in hIAPP fibrillation. The negatively charged carboxylate groups of pyromellitic acid moiety of C5 displayed interactions with the key residues of hIAPP as noted in the conformational clustering analysis. Notably higher sampling of helix from 24.00 ± 0.84 % in hIAPP to 34.20 ± 1.92 % in hIAPP–C5 is consistent with the CD studies, which depicted that C5 trapped hIAPP monomer in a conformation that blocked its self-association. MD simulations illuminated the molecular mechanism and atomistic details of the C5 interactions with hIAPP, which are responsible for its inhibitory activity against hIAPP fibrillation and alleviating hIAPP aggregates-induced cytotoxicity.