Role of Site-Specific Asparagine Deamidation in Islet Amyloid Polypeptide Amyloidogenesis: Key Contributions of Residues 14 and 21

5 years ago

Role of Site-Specific Asparagine Deamidation in Islet Amyloid Polypeptide Amyloidogenesis: Key Contributions of Residues 14 and 21

Ximena Zottig, Phuong Trang Nguyen, Steve Bourgault, Mathew Sebastiao

Deamidation of an asparagine residue is a spontaneous non-enzymatic post-translational modification that results in the conversion of asparagine into a mixture of aspartic acid and isoaspartic acid. This chemical conversion modulates protein conformation and physicochemical properties, which could lead to protein misfolding and aggregation. In this study, we investigated the effects of site-specific Asn deamidation on the amyloidogenicity of the aggregation-prone peptide islet amyloid polypeptide (IAPP). IAPP is a 37-residue peptidic hormone whose deposition as insoluble amyloid fibrils is closely associated with type 2 diabetes. Asn residues were successively substituted with an Asp or isoAsp, and amyloid formation was evaluated by a thioflavin T fluorescence assay, circular dichroism spectroscopy, atomic force microscopy, and transmission electron microscopy. Whereas deamidation at position 21 inhibited IAPP conformational conversion and amyloid formation, the N14D mutation accelerated self-assembly and led to the formation of long and thick amyloid fibrils. In contrast, IAPP was somewhat tolerant to the successive deamidation of Asn residues 22, 31, and 35. Interestingly, a small molar ratio of IAPP deamidated at position 14 promoted the formation of nucleating species and the elongation from unmodified IAPP. Besides, using the rat pancreatic β cell line INS-1E, we observed that site-specific deamidation did not significantly alter IAPP-induced toxicity. These data indicate that Asn deamidation can modulate IAPP amyloid formation and fibril morphology and that the site of modification plays a critical role. Above all, this study reinforces the notion that IAPP amyloidogenesis is governed by precise intermolecular interactions involving specific Asn side chains.

Publisher URL: http://dx.doi.org/10.1021/acs.biochem.7b00209

DOI: 10.1021/acs.biochem.7b00209