G protein-coupled receptors (GPCRs) are involved in a wide variety of physiological processes. Therefore, approximately 40% of currently prescribed drugs have targeted this receptor family. Discovery of β -arrestin mediated signaling and also separability of G protein and β -arrestin signaling pathways have switched the research focus in the GPCR field towards development of biased ligands, which provide engagement of the receptor with a certain effector, thus enriching a specific signaling pathway. In this review, we summarize possible factors that impact signaling profiles of GPCRs such as oligomerization, drug treatment, disease conditions, genetic background, etc. along with relevant molecules that can be used to modulate signaling properties of GPCRs such as allosteric or bitopic ligands, ions, aptamers and pepducins. Moreover, we also discuss the importance of inclusion of pharmacogenomics and molecular dynamics simulations to achieve a holistic understanding of the relation between genetic background and structure and function of GPCRs and GPCR-related proteins. Consequently, specific downstream signaling pathways can be enriched while those that bring unwanted side effects can be prevented on a patient-specific basis. This will improve studies that centered on development of safer and personalized therapeutics, thus alleviating the burden on economy and public health.

This is an open access article under the terms of the Creat ive Commo ns Attri bution-NonCo mmercial License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited and is not used for commercial purposes.

Coronavirus disease-2019 (COVID-19) was firstly reported in Wuhan, China, towards the end of 2019, and emerged as a pandemic. The spread and lethality rates of the COVID-19 have ignited studies that focus on the development of therapeutics for either treatment or prophylaxis purposes. In parallel, drug repurposing studies have also come into prominence. Herein, we aimed at having a holistic understanding of conformational and dynamical changes induced by an experimentally characterized inhibitor on main protease (M pro ) which would enable the discovery of novel inhibitors. To this end, we performed molecular dynamics simulations using crystal structures of apo and α -ketoamide 13b-bound M pro homodimer. Analysis of trajectories pertaining to apo M pro revealed a new target site, which is located at the homodimer interface, next to the catalytic dyad. Thereafter, we performed ensemble-based virtual screening by exploiting the ZINC and DrugBank databases and identified three candidate molecules, namely eluxadoline, diosmin, and ZINC02948810 that could invoke local and global conformational rearrangements which were also elicited by α -ketoamide 13b on the catalytic dyad of M pro. Furthermore, ZINC23881687 stably interacted with catalytically important residues Glu166 and Ser1 and the target site throughout the simulation. However, it gave positive binding energy, presumably, due to displaying higher flexibility that might dominate the entropic term, which is not included in the MM-PBSA method. Finally, ZINC20425029, whose mode of action was different, modulated dynamical properties of catalytically important residue, Ala285. As such, this study presents valuable findings that might be used in the development of novel therapeutics against M pro. Communicated by Ramaswamy H. Sarma

Undruggability of RAS proteins has necessitated alternative strategies for the development of effective inhibitors. In this respect, phosphorylation has recently come into prominence as this reversible post-translational modification attenuates sensitivity of RAS towards RAF. As such, in this study, we set out to unveil the impact of phosphorylation on dynamics of HRASWT and aim to invoke similar behavior in HRASG12D mutant by means of small therapeutic molecules. To this end, we performed molecular dynamics (MD) simulations using phosphorylated HRAS and showed that phosphorylation of Y32 distorted Switch I, hence the RAS/RAF interface. Consequently, we targeted Switch I in HRASG12D by means of approved therapeutic molecules and showed that the ligands enabled detachment of Switch I from the nucleotide-binding pocket. Moreover, we demonstrated that displacement of Switch I from the nucleotide-binding pocket was energetically more favorable in the presence of the ligand. Importantly, we verified computational findings in vitro where HRASG12D/RAF interaction was prevented by the ligand in HEK293T cells that expressed HRASG12D mutant protein. Therefore, these findings suggest that targeting Switch I, hence making Y32 accessible might open up new avenues in future drug discovery strategies that target mutant RAS proteins.

RAS mutants have been extensively studied as they are associated with development of cancer; however, H-RAS G12P mutant has remained untouched since it does not lead to transformation in the cell. To the best of our knowledge, this is the first study where structural/dynamical properties of H-RAS G12P have been investigated -in comparison to H-RAS WT , H-RAS G12D , RAF-RBD-bound and GAP-bound H-RAS WT - using molecular dynamics simulations (total of 9 μ s). We observed remarkable differences in dynamics of Y32. Specifically, it is located far from the nucleotide binding pocket in the catalytically-active GAP-bound H-RAS WT , whereas it makes close interaction with the nucleotide in signaling-active systems (H-RAS G12D , KRAS4B G12D , RAF-RBD-bound H-RAS WT ) and H-RAS WT . The accessibility of Y32 in wild type protein is achieved upon GAP binding. Interestingly; however, it is intrinsically accessible in H-RAS G12P . Considering the fact that incomplete opening of Y32 is associated with cancer, we propose that Y32 can be targeted by means of small therapeutics that can displace it from the nucleotide binding site, thus introducing intrinsic GTPase activity to RAS mutants, which cannot bind to GAP. Therefore, mimicking properties of H-RAS G12P in RAS-centered drug discovery studies has the potential of improving success rates since it acts as a molecular switch per se .