ELUCIDATING THE MOLECULAR MECHANISMS OF AGGREGATION OF FUNCTIONAL AMYLOIDS FORMED BY HUMAN NEUROPEPTIDE Y

Amyloids are a type of protein aggregate that adopts a highly ordered and stable fibrillar structure. They are associated with various neurodegenerative diseases, such as Alzheimer`s and Parkinson`s, as well as other systemic disorders. Amyloids form when certain proteins undergo a misfolding process, leading to their aggregation into insoluble, beta-sheet-rich structures. These aggregates accumulate in tissues and organs, impairing their normal function and causing cellular toxicity. The exact role of amyloids in disease pathogenesis is not fully understood, but evidence suggests they can disrupt cellular processes, induce inflammation, and cause cell death. Studying the molecular mechanisms of amyloid formation is essential for gaining insights into disease progression and developing potential therapeutic strategies to target and prevent amyloid-related disorders. This project focuses on the study of the molecular mechanisms of functional amyloid formation, with a focus on neuropeptides like Neuropeptide Y. These hormones, which play a crucial role in modulating neuronal activity and regulating a variety of physiological processes, are stored and released in the form of amyloids in nerve cells. Despite the nefarious role of amyloid aggregates in the evolution of neurodegenerative diseases, they also perform essential biological functions, forming an emerging study area. We have developed a refined protocol for efficiently producing purified Neuropeptide Y, utilizing heterologous expression in E. coli BL21 (DE3) Star strains. A range of E. coli growth and protein expression strategies, including the application of an auto-induction medium, have been evaluated. As part of our ongoing project, we are conducting high-throughput experiments on amyloid aggregation kinetics using a microplate fluorescence detection system. These experiments are accompanied by the identification of beta-rich structure formation, characteristic of amyloid structures, via circular dichroism. With this project, we seek to elucidate the aggregation and disaggregation kinetics of a functional amyloid example and investigate the mechanisms that allow cells to control this process without generating pathogenic structures. Understanding these processes could provide new perspectives for treating neurodegenerative diseases and expand our understanding of stress physiology and regulation of the nervous system.