A2A Adenosine Receptor Partial Agonism Related to Structural Rearrangements in an Activation Microswitch

The role of structural dynamics in GPCR‐mediated signaling

“…While no detailed structural information is available for a partial agonist-bound intermediate state of the receptor, the chemical shift differences between the substates of the fluorine-labeled TM6 of the A 2A R in 19 F-NMR spectra indicate that partial agonists increase the population of a receptor conformation with a less open intracellular transducer-binding cavity, in agreement with the previously mentioned studies on the b 2 AR [78,117,119]. This is in line with a very recent [ 15 N, 1 H]-TROSY NMR study that revealed differences in the local structure at the intracellular surface of TM6 of the A 2A R between full and partial agonists [84]. Taken together, these results are consistent with the model that partial agonists preferentially stabilize conformations, which are distinct and less efficacious from the ones that are promoted by full agonists, rather than not being able to completely shift the equilibrium toward the active receptor conformation.…”

“…[58]), electron paramagnetic resonance (EPR) (reviewed in Refs [59,60]), and fluorescence spectroscopy (reviewed in Refs [61,62]) supported by molecular dynamics (MD) simulations (reviewed in Refs [63,64]) have been used in complementation to crystallographic and cryo-EM studies to analyze different conformational states of GPCRs and to determine their liganddependent energetics and rates of interconversion. Several NMR studies on the b 1 and b 2 adrenergic receptors (b 1 AR and b 2 AR) [65][66][67][68][69][70][71][72][73][74][75][76][77][78][79][80], the adenosine A 2A receptor (A 2A R) [81][82][83][84][85][86][87], the l-opioid receptor (lOR) [88,89], the leukotriene B 4 receptor 2 (BLT2R) [90], the a 1A adrenergic receptor (a 1A R) [91], the neurotensin receptor type 1 (NTSR1) [92], and the M 2 R [93,94] have shown that GPCRs are highly dynamic proteins that exist in an equilibrium between multiple functionally relevant conformational states. Among these receptors, the b 2 AR and the A 2A R are the most extensively studied GPCRs in terms of their conformational dynamics.…”

“…Recent NMR studies using [ 15 N, 1 H]-TROSY measurements on the A 2A R provided further evidence for the importance of rearrangements in the PIF motif for receptor activation as well as the role of the conserved residue D52 2.50 of the sodium-binding site as an allosteric switch [82,84]. Analysis of chemical shift changes of the well-separated tryptophane indole and glycine backbone NMR signals in the presence of antagonists, partial agonists, and full agonists revealed different conformations in the receptor core that correlated with the efficacy of the bound ligand [84]. In particular, the chemical shift changes of the toggle switch residue W246 6.48 , that has been proposed to report on differences in the relative orientation of F242 6.44 , suggest that the PIF motif samples distinct conformations in the presence of antagonists, partial agonists, and full agonists, respectively.…”

“…Recent NMR studies using [ 15 N, 1 H]‐TROSY measurements on the A 2A R provided further evidence for the importance of rearrangements in the PIF motif for receptor activation as well as the role of the conserved residue D52 2.50 of the sodium‐binding site as an allosteric switch [82,84]. Analysis of chemical shift changes of the well‐separated tryptophane indole and glycine backbone NMR signals in the presence of antagonists, partial agonists, and full agonists revealed different conformations in the receptor core that correlated with the efficacy of the bound ligand [84].…”

Production of a Human Histamine Receptor for NMR Spectroscopy in Aqueous Solutions

“…While several dozen GPCRs have been reported to be produced in P. pastoris [ 11 ], including adrenergic receptors [ 12 ], muscarinic receptors [ 13 ], dopamine receptors [ 14 ] and opioid receptors [ 15 ], a much smaller number of GPCRs have been successfully produced in P. pastoris for NMR experiments. The most studied example is the human A 2A adenosine receptor [ 16 , 17 , 18 , 19 , 20 , 21 , 22 , 23 ]. More recently, NMR data have been reported for the cannabinoid receptor type 1 [ 24 ] and orexin receptor 2 [ 24 ] proteins expressed in P. pastoris .…”