Alisha D. Caliman

The sphingolipid S1P allosterically activates atypical protein kinase C to promote cell survival.

The A_2A adenosine receptor (A_2AAR) is a G protein‐coupled receptor that is pharmacologically targeted for the treatment of inflammation, sepsis, cancer, neurodegeneration, and Parkinson's disease. Recently, we applied long‐timescale molecular dynamics simulations on two ligand‐free receptor conformations, starting from the agonist‐bound (PDB ID: 3QAK) and antagonist‐bound (PDB ID: 3EML) X‐ray structures. This analysis revealed four distinct conformers of the A_2AAR: the active, intermediate 1, intermediate 2, and inactive. In this study, we apply the fragment‐based mapping algorithm, FTMap, on these receptor conformations to uncover five non‐orthosteric sites on the A_2AAR. Two sites that are identified in the active conformation are located in the intracellular region of the transmembrane helices (TM) 3/TM4 and the G protein‐binding site in the intracellular region between TM2/TM3/TM6/TM7. Three sites are identified in the intermediate 1 and intermediate 2 conformations, annexing a site in the lipid interface of TM5/TM6. Five sites are identified in the inactive conformation, comprising a site in the intracellular region of TM1/TM7 and in the extracellular region of TM3/TM4 of the A_2AAR. We postulate that these sites on the A_2AAR be screened for allosteric modulators for the treatment of inflammatory and neurological diseases.

Activation of the first sphingosine-1-phosphate receptor (S1PR₁) promotes permeability of the blood brain barrier, astrocyte and neuronal protection, and lymphocyte egress from secondary lymphoid tissues. Although an agonist often activates the S1PR₁, the receptor exhibits high levels of basal activity. In this study, we performed long-timescale molecular dynamics and accelerated molecular dynamics (aMD) simulations to investigate activation mechanisms of the ligand-free (apo) S1PR₁. In the aMD enhanced sampling simulations, we observed four independent events of activation, which is characterized by close interaction between Y311^7.53 and Y221^5.58 and increased distance between the intracellular ends of transmembrane (TM) helices 3 and 6. Although transmembrane helices TM3, TM6, TM5 and, TM7 are associated with GPCR activation, we discovered that their movements are not necessarily correlated during activation. Instead, TM5 showed a decreased correlation with each of these regions during activation. During activation of the apo receptor, Y221^5.58 and Y311^7.53 became more solvated, because a water channel formed in the intracellular pocket. Additionally, a lipid molecule repeatedly entered the receptor between the extracellular ends of TM1 and TM7, providing important insights into the pathway of ligand entry into the S1PR₁.

The activation/deactivation processes for G-protein coupled receptors (GPCRs) have been computationally studied for several different classes, including rhodopsin, the β2 adrenergic receptor, and the M2 muscarinic receptor. Despite determined cocrystal structures of the adenosine A_2A receptor (A_2AAR) in complex with antagonists, agonists and an antibody, the deactivation process of this GPCR is not completely understood. In this study, we investigate the convergence of two apo simulations, one starting with an agonist-bound conformation (PDB: 3QAK)¹⁴ and the other starting with an antagonist-bound conformation (PDB: 3EML)¹¹. Despite the two simulations not completely converging, we were able to identify distinct intermediate steps of the deactivation process characterized by the movement of Y288^7.53 in the NPxxY motif. We find that Y288^7.53 contributes to the process by forming hydrogen bonds to residues in transmembrane helices 2 and 7 and losing these interactions upon full deactivation. Y197^5.58 also plays a role in the process by forming a hydrogen bond only once the side chain moves from the lipid interface to the middle of the helical bundle.