Unveiling Metastable Ensembles of GRB2 and the Relevance of Interdomain Communication during Folding

Autor(es)

Dias, Raphael V. R. ; Pedro, Renan P. ; Sanches, Murilo N. ; Moreira, Giovana C. ; Leite, Vitor B. P. ; Caruso, Icaro P. ; de Melo, Fernando A. ; de Oliveira, Leandro C.

• Tipo de documento: Artigo Científico
• Fonte: JOURNAL OF CHEMICAL INFORMATION AND MODELING; v. 63, n. 20, p. 10-pg., 2023-10-12.

Resumo

The folding process of multidomain proteins is a highly intricate phenomenon involving the assembly of distinct domains into a functional three-dimensional structure. During this process, each domain may fold independently while interacting with others. The folding of multidomain proteins can be influenced by various factors, including their composition, the structure of each domain, or the presence of disordered regions, as well as the surrounding environment. Misfolding of multidomain proteins can lead to the formation of nonfunctional structures associated with a range of diseases, including cancers or neurodegenerative disorders. Understanding this process is an important step for many biophysical analyses such as stability, interaction, malfunctioning, and rational drug design. One such multidomain protein is growth factor receptor-bound protein 2 (GRB2), an adaptor protein that is essential in regulating cell survival. GRB2 consists of one central Src homology 2 (SH2) domain flanked by two Src homology 3 (SH3) domains. The SH2 domain interacts with phosphotyrosine regions in other proteins, while the SH3 domains recognize proline-rich regions on protein partners during cell signaling. Here, we combined computational and experimental techniques to investigate the folding process of GRB2. Through computational simulations, we sampled the conformational space and mapped the mechanisms involved by the free energy profiles, which may indicate possible intermediate states. From the molecular dynamics trajectories, we used the energy landscape visualization method (ELViM), which allowed us to visualize a three-dimensional (3D) representation of the overall energy surface. We identified two possible parallel folding routes that cannot be seen in a one-dimensional analysis, with one occurring more frequently during folding. Supporting these results, we used differential scanning calorimetry (DSC) and fluorescence spectroscopy techniques to confirm these intermediate states in vitro. Finally, we analyzed the deletion of domains to compare our model outputs to previously published results, supporting the presence of interdomain modulation. Overall, our study highlights the significance of interdomain communication within the GRB2 protein and its impact on the formation, stability, and structural plasticity of the protein, which are crucial for its interaction with other proteins in key signaling pathways. (AU)

• Processo FAPESP: 19/24974-0 - Estudos funcionais da proteína growth-factor-receptor bound protein 2 (GRB2) por meio de ressonância magnética nuclear e fluorescência: correlação entre dinâmica e estrutura
• Beneficiário: Fernando Alves de Melo
• Modalidade de apoio: Auxílio à Pesquisa - Regular
• Processo FAPESP: 14/17630-0 - Estudos funcionais da tirosina quinase Fibroblast Growth Factor receptor (FGFR2), da adaptadora growth factor Receptor- bound protein 2 (GRB2) e da tirosina fosfatase SHP2
• Beneficiário: Fernando Alves de Melo
• Modalidade de apoio: Auxílio à Pesquisa - Regular
• Processo FAPESP: 21/15028-4 - Elucidando a plasticidade funcional de proteínas intrinsecamente desordenadas pelo estudo de relevo de energia
• Beneficiário: Murilo Nogueira Sanches
• Modalidade de apoio: Bolsas no Brasil - Doutorado
• Processo FAPESP: 19/22540-3 - Estudos de relevo de superfícies de energia de macromoléculas biológicas
• Beneficiário: Vitor Barbanti Pereira Leite
• Modalidade de apoio: Auxílio à Pesquisa - Regular
• Processo FAPESP: 23/02219-1 - Entendendo mecanismos funcionais de proteínas e RNAs por meio de relevos de superfícies de energia
• Beneficiário: Vitor Barbanti Pereira Leite
• Modalidade de apoio: Auxílio à Pesquisa - Regular
• Processo FAPESP: 16/08753-6 - Avaliação de mecanismos de quinases e proteínas relacionadas
• Beneficiário: Raphael Vinicius Rodrigues Dias
• Modalidade de apoio: Bolsas no Brasil - Mestrado