Investigating by circular dichroism some amyloidogenic elastin-derived polypeptides.

Tamburro and coworkers have demonstrated that some elastin-derived polypeptide sequences are able to give rise, in vitro, to amyloid-like fibers. The biological relevance of this finding could be explained by the recent detection of some amyloidogenic material found in arteries of old patients affected by atherosclerosis and demonstrated to be elastin derived. In this context, the comprehension of the mechanism responsible for the amyloid-like fibrillogenesis of elastin-derived sequences is of crucial importance for the design of drugs that could inhibit the amyloidogenic process. To gain further insights into the elastin amyloidogenic process, we studied the polypeptide sequences encoded by Exon 7 and Exon 32 of the human tropoelastin gene, and we demonstrated that only Exon 32 is able to aggregate in amyloid-like fibers. Vis-UV Thioflavin T circular dichroism (CD) spectroscopy rapidly and unambiguously detected the amyloidogenic propensity of the polypeptides. To gain additional insights into the aggregation mechanism of elastin-derived amyloidogenic peptides, we carried out the kinetics of EX32 amyloid-like aggregates by using ThT dye. CD spectroscopy was also used for investigating the secondary structure of the polypeptides, thus giving useful insights into the conformations involved in amyloid-like fiber formation. Furthermore, complementary techniques such as fluorescence spectroscopy, spectral shift, and binding Congo red UV assays as well as atomic force microscopy were also used to confirm the amyloidogenic behavior of the studied polypeptides.