FRPFI

Abstract

Correct protein folding is pivotal for normal cell physiology and its failure is related to the development of several human diseases, including neurodegeneration and cancer. Over the last decades, the problem of characterizing the protein folding process has been mostly tackled with the aim of clarifying the physico-chemical mechanisms involved in the exploration of the energy landscape and in the efficient recognition of the native structure. Biomedical implications of understanding the protein folding pathways have been mostly associated with misfolding and amyloidogenic aggregation processes. Very recently, the Trento unit has conceived and validated a new paradigm for rational drug discovery, based on identifying small molecules that bind to pockets in the metastable conformations visited along the protein folding pathway and stabilize them, thus triggering the degradation pathways. This approach is enabled by some powerful computational schemes for protein folding simulations developed in the Trento unit, which make it possible to simulate the folding of biologically relevant proteins using realistic all-atom force fields, given their three-dimensional native structure. The resulting drug discovery scheme has already been successfully applied to several target proteins, in both academic and industrial research. In this project, we take inspiration from this translational research and address a related fundamental question: do protein folding intermediates also play a biological role? Specifically, we aim at clarifying if intermediates represent an unappreciated layer of physiological regulation of protein expression, through post-translational modifications. We shall focus on two specific proteins (Androgen Receptor (AR) and von Hippel-Lindau tumor suppressor (pVHL)) that are known to be post-translationally regulated via phosphorylation. In these proteins, phosphorylation sites that play a role their homeostasis regulation are deeply buried into the core of the native structure, thus are inaccessible to kinases. To test if the phosphorylation may occur along the folding pathway, we will adopt a highly integrated cross-disciplinary approach, combining data mining, all-atom molecular simulations, biochemical analyses, and biophysical experiments. Indeed, the Trento unit will employ its cutting-edge technology for all-atom protein folding simulations, while the Padova unit will take advantages of established competencies in bioinformatics and cell biochemistry, along with direct research experience on the selected proteins, and the Milano Unit will exploit single-molecule force spectroscopy experiments to study the structural dynamics of biomolecules. The proof of a functional role for protein folding intermediates in post translational regulation would have a tremendous impact in life science, unveiling an entirely new regulation layer in the Fundamental Dogma of Biology, with a wide range of potential biomedical implications.

Progetto PRIN 2022TWRXAW funded by the Italian Ministry of University and Research.