ELUCIDATING THE MOLECULAR MECHANISMS OF AGGREGATION OF FUNCTIONAL AMYLOID FORMED BY HUMAN CORTICOTROPIN-RELEASING HORMONE

Amyloids are a type of protein aggregate that adopts a highly ordered and stable fibrillar beta-sheet-rich structure. The study of fibril formation is a key area of research due to its association with several neurodegenerative diseases, as well as the unique ability of several proteins to form these structures, even without shared sequence similarities. The exact role of amyloids in neurodegenerative diseases is not fully understood, but evidence suggests they can disrupt cellular processes, induce inflammation, and cause cell death.

This project aims to elucidate the kinetics of aggregation and disaggregation of an important specimen of functional amyloid, the Corticotropin-releasing hormone (CRH). We will investigate the mechanisms that allow cells to control this process without generating pathogenic structures. CRH is fundamental in the body's response to stress, with elevated levels associated with depression and significantly higher in the cerebrospinal fluid of individuals who have committed suicide.

We are developing a refined protocol to efficiently produce and purify human CRH using heterologous expression in E. coli BL21 (DE3) Star strains. As part of our ongoing project, we can identify and characterize the formation of amyloid fibers through the presence of beta-rich structures using circular dichroism. To better understand the mechanisms of amyloid aggregation kinetics, we use a system to detect the fibers formation via fluorescence in microplates. The obtained data set allows for a global fitting to determine which fibril formation model best fits the data Understanding these processes could provide new perspectives for treating neurodegenerative diseases and broaden our understanding of stress physiology and the regulation of the nervous system.