Grb2 is an important regulator of normal vs. oncogenic cell signaling transduction. It plays a pivotal role on kinase-mediated signaling transduction by linking Receptor Tyrosine kinases to Ras/MAPK pathway which is known to bring oncogenic outcome. Coumarins are phenolic molecules found in several plants and seeds widely studied because of the antibiotic, anti-inflammatory, anticoagulant, vasodilator, and anti-tumor properties. Despite several studies about the anti-tumor properties of Coumarin in vivo and the role of Grb2 in signaling pathways related to cell proliferation, a molecular level investigation of the interaction between Grb2 and Coumarin is still missing. In this study, we performed a combined set of biophysical approaches to get insights on the interaction between Grb2 in a dimer state and Coumarin. Our results showed that Coumarin interacts with Grb2 dimer through its SH2 domain. The interaction is entropically driven, 1:1 molecular ratio and presents equilibrium constant of 105 M-1. In fact, SH2 is a well-known domain and a versatile signaling module for drug targeting which has been reported to bind compounds that block Ras activation in vivo. Despite we don't know the biological role coming from interaction between Grb2-SH2 domain and Coumarin, it is clear that this molecule could work in the same way as a SH2 domain inhibitor in order to block the link of Receptor Tyrosine kinases to Ras/MAPK pathway.

The human Respiratory Syncytial Virus (hRSV) stands as one of the most common causes of acute respiratory diseases. The infectivity of this virus is intricately linked to its membrane proteins, notably the attachment glycoprotein (G protein). The latter plays a key role in facilitating the attachment of hRSV to respiratory tract epithelial cells, thereby initiating the infection process. The present study aimed to characterize the interaction of the conserved cysteine-noose domain of hRSV G protein (cndG) with the transmembrane CX3C motif chemokine receptor 1 (CX3CR1) isoforms using computational tools of molecular modeling, docking, molecular dynamics simulations, and binding free energy calculations. From MD simulations of the molecular system embedded in the POPC lipid bilayer, we showed a stable interaction of cndG with the canonical fractalkine binding site in the N-terminal cavity of the CX3CR1 isoforms and identified that residues in the extracellular loop 2 (ECL2) region and Glu279 of this receptor are pivotal for the stabilization of CX3CR1/cndG binding, corroborating what was reported for the interaction of the chemokine fractalkine with CX3CR1 and its structure homolog US28. Therefore, the results presented here contribute by revealing key structural points for the CX3CR1/G interaction, allowing us to better understand the biology of hRSV from its attachment process and to develop new strategies to combat it.

The respiratory syncytial virus (RSV) matrix protein plays key roles in the virus life cycle and is essential for budding, as it stimulates the optimal membrane curvature necessary for the emergence of viral particles. Resveratrol, a polyphenol (3,4′,5-trihydroxy-trans-stilbene) produced by plants, exhibits pharmacological effects, including anti-inflammatory and antiviral activities. In this study, resveratrol was tested in HEp-2 (Epidermoid carcinoma of the larynx cell) cells for its post-infection effects, and recombinant M protein was produced to characterize the biophysical mechanisms underlying this interaction. The CC50 (Cytotoxic concentration 50%) value for resveratrol was determined to be 297 μM over 48 h, and the results from the HEp-2 cell cultures demonstrated a viral inhibition of 42.7% in the presence of resveratrol, with an EC50 (Half maximal effective concentration) of 44.26 μM. This mechanism may occur through interaction with the M protein responsible for the budding of mature viral particles. Biophysical assays enabled us to characterize the interaction of the M/resveratrol complex as an entropically driven bond, guided by hydrophobic interactions at the dimerization interface of the M protein, which is essential for the stabilization and formation of the oligomers necessary for viral budding. These findings suggest that one of the targets for resveratrol binding is the M protein, indicating a potential site for blocking the progression of the infection.

The Growth Factor Receptor-Bound Protein 2 (Grb2) is a central adaptor protein in signal transduction pathways, yet how its monomer-dimer equilibrium governs its supramolecular organization remains elusive. Here, we demonstrate that the oligomeric state of Grb2 acts as a binary switch for Liquid-Liquid Phase Separation (LLPS). While the auto-inhibited homodimer forms only transient, thermodynamically unstable assemblies in crowding conditions, the phosphorylation-mimetic monomer (Y160E) drives the formation of robust, gel-like condensates. Integrating turbidity assays, temperature-controlled dynamic light scattering, fluorescence recovery after photobleaching (FRAP), hyperspectral imaging analyses, and coarse-grained molecular dynamics simulations, we reveal that this phase transition is enthalpy-driven and reliant on a specific electrostatic network between the SH2 domain residue R142 and a C-terminal SH3 acidic cluster (Q170-D172). Crucially, we show that these monomeric condensates function as "scaffolds" that efficiently recruit and sequester cytosolic wild-type dimers ("clients") into the dense phase. This recruitment mechanism resolves the paradox of how non-condensing wild-type proteins participate in phase separation. Our findings propose a novel regulatory model where phosphorylation nucleates the formation of high-density signaling hubs, redefining Grb2 from a passive adaptor to a dynamic spatial organizer of the Ras/MAPK pathway.

The dysregulated activation of the Ras pathway is directly associated with approximately one-third of human cancer cases. The interaction between the adaptor protein GRB2 and the guanine nucleotide exchange factor SOS1 plays a crucial role in this process, serving as a key link in the activation of Ras. This interaction facilitates signal transduction that regulates essential cellular processes such as proliferation, survival, and differentiation and is, therefore, a central point in oncogenesis. Although the GRB2-SOS1 complex is critical for the regulation of Ras pathway signaling, the structural mechanisms governing this interaction remain largely unknown. In this study, we conducted an in-depth analysis of the interaction between GRB2 and peptides derived from SOS1, utilizing computational modeling, docking, and molecular dynamics simulations, combined with a detailed analysis of the energy landscape (ELViM). The results demonstrate that the conformational flexibility of the GRB2 protein has a direct impact on the stability of the GRB2-SOS1 complex, with communication between GRB2 domains being a determining factor for the robustness of this interaction. Additionally, we identified critical residues that play decisive roles in the formation and regulation of this interaction, which may serve as potential targets for modulation of the Ras pathway. These findings not only deepen the understanding of the structural mechanisms underlying the GRB2–SOS1 interaction but also provide a foundation for the development of specific therapeutic strategies aimed at controlling aberrant Ras pathway signaling, thereby offering new prospects in the treatment of diseases associated with the anomalous activation of Ras.