Correlation between the structure and dynamics of GRB2 protein and its folding excited states

Abstract

Cellular metabolism is mediated by signaling pathways that are regulated by enzyme activities dependent on the reversible phosphorylation of the amino acid side chain. In this way, tyrosine kinase proteins (PTKs) and tyrosine phosphatase proteins (PTPs) are pivotal in the regulation of metabolism, gene expression, growth, division and cellular differentiation. Once phosphorylated, PTKs recruit partner proteins (Grb2, Shc, Ras, Sos, etc.) to form primary signaling complexes (ESCs), which activate specific signaling pathways within the cell. In many cases, kinases and phosphatases act together in these processes. In this context, FGFR2 and Shp2 are "key" PTKs and PTPs of many of the signaling processes whose aberrant activity causes a variety of cancers and fetal malformations. Recent studies show that the protein Grb2 (Growth-factor receptor bound protein 2) is an important regulator of FGFR2, preventing its kinase activity before extracellular stimulation. When phosphorylated, Grb2 dissociates from FGFR2, resulting in the initiation of its enzymatic activity that results in the recruitment of partner proteins in the signaling cascade. De-phosphorylation of Grb2 by Shp2 rescues the previously formed complex, re-imposing control over FGFR2. Another recent study claims that Grb2 regulates the activity of the MAPK signaling pathway (Mitogen-activated protein kinases). According to this study, only in its monomeric form, Grb2 would be able to bind to SOS (guanine nucleotide exchange factors) and activate the MAPK pathway, while in its dimeric form, Grb2 would be an inhibitor of this process. Grb2 is therefore a global controller of these mutually dependent reactions that are related to the development of various human dysplasia, and therefore an important biological target due to its versatility (in addition to being an adaptor protein) in performing functions within the cell. Thus, we intend to use Molecular Dynamics, CPMG (Carr-Purcell-Meiboom-Gill), and CEST (Chemical Exchange Saturation Transfer) to study Grb2 dimer and monomer, in order to access their dynamic modes under different physical-chemical conditions, excited states and unfolding routes, and conformational changes induced by phosphorylation and/or ligands with antitumor properties. The characterization of these multifunctional properties of Grb2, which is inherent in flexible, multidomain and intrinsically disordered proteins, is fundamental for understanding the versatility of functions within cells associated with this protein and associated with cellular proliferation. (AU)