Aromatic Interactions Are Not Required for Amyloid Fibril Formation by Islet Amyloid Polypeptide but Do Influence the Rate of Fibril Formation and Fibril Morphology

Abstract

Amyloid formation has been implicated in a wide range of human diseases, and a diverse set of proteins is involved. There is considerable interest in elucidating the interactions which lead to amyloid formation and which contribute to amyloid fibril stability. Recent attention has been focused upon the potential role of aromatic-aromatic and aromatic-hydrophobic interactions in amyloid formation by short to midsized polypeptides. Here we examine whether aromatic residues are necessary for amyloid formation by islet amyloid polypeptide (IAPP). IAPP is responsible for the formation of islet amyloid in type II diabetes which is thought to play a role in the pathology of the disease. IAPP is 37 residues in length and contains three aromatic residues, Phe-15, Phe-23, and Tyr-37. Structural models of IAPP amyloid fibrils postulate that Tyr-37 is near one of the phenylalanine residues, and it is known that Tyr-37 interacts with one of the phenylalanines during fibrillization; however, it is not known if aromatic-aromatic or aromatic-hydrophobic interactions are absolutely required for amyloid formation. An F15L/F23L/Y37L triple mutant (IAPP-3XL) was prepared, and its ability to form amyloid was tested. CD, thioflavin binding assays, AFM, and TEM measurements all show that the triple leucine mutant readily forms amyloid fibrils. The substitutions do, however, decrease the rate of fibril formation and alter the tendency of fibrils to aggregate. Thus, while aromatic residues are not an absolute requirement for amyloid formation by IAPP, they do play a role in the fibril assembly process.

Aliphatic peptides show similar self-assembly to amyloid core sequences, challenging the importance of aromatic interactions in amyloidosis

“…Our multimodal analysis of the amyloid peptide aggregates clearly indicates that aromatic residues are not essential for self-assembly. Nevertheless, they alter the kinetics of aggregation as seen previously with the whole amyloid protein (35) and increase the proportion of disordered aggregates. Molecular dynamics simulations show that NL 6 and KE 7 form more disordered aggregates compared with LD 6 and ID 3 in the initial stages.…”

Islet amyloid polypeptide toxicity and membrane interactions

“…This peptide is not toxic and is a potent inhibitor of amyloid formation and cytotoxicity by wild-type hIAPP (34). Fifth, the 3XL-IAPP peptide is a triple mutant of hIAPP in which the three aromatic residues, F15, F23, and Y37, have been replaced by L (35). It forms amyloid approximately ninefold more slowly than wild-type hIAPP and allows us to test the role of the aromatic residues in membrane disruption.…”

“…The triple leucine mutant, 3XL-IAPP, was originally designed to test the role of the aromatic residues in IAPP amyloid formation (35). 3XL-IAPP forms amyloid fibrils with a similar morphology to those derived from wild-type hIAPP, but does so more slowly.…”

Converting the Highly Amyloidogenic Human Calcitonin into a Powerful Fibril Inhibitor by Three-dimensional Structure Homology with a Non-amyloidogenic Analogue

“…We have suggested that, for an essentially unstructured polypeptide chain like hCT, orientation could be driven by a hydrogen bond donor and the center of a benzene ring, which acts as a hydrogen bond acceptor (20,61). The relevance of aromatic residues was postulated (62), predicted theoretically (63), and shown experimentally (64) to increase the rate of fibril formation. Accordingly, intermolecular hydrogen bonds have been reported to play an important role in the association of the hCT molecules (58).…”