The Adenosine A2a Receptor

The T4L fusion strategy in combination with the lipidic cubic phase crystallization method that resulted in the structure of the p2AR has also been applied successfully to the adenosine A2a receptor. This suggests that the T4L technology is transferable across different subfamilies of Class A GPCRs. In this structure, obtained to 2.6 A resolution, the receptor is crystallized in complex with the high-affinity antagonist ZM241385. Incorporation of the T4L moiety shifts the conformation to an agonist-like state as defined by an increase in the affinity of agonist and no change in antagonist affinity.

The adenosine A2a-T4L structure had a number of features not seen in previous structures. Although the overall arrangement of the helices was

Figure 14.8 The binding site of the antagonist ZM241385 (pink) in the adenosine A2a receptor showing its vertical position within the A2a binding pocket.

similar to the other structures, a change in the relative positions of the helices results in a movement of the antagonist binding site. TM6 and TM7 are shifted closer to the binding pocket by 7 A and 3 A, respectively, and there is a lateral shift in TM3 toward TM5 by 3 A . In contrast to carazolol and cyanopindolol, the antagonists for the P-receptors, ZM241385 lies in a plane perpendicular to the membrane (Fig. 14.8). The bicyclic core of this compound interacts with the aromatic ring of Phe168529 in the second extracellular loop, while Trp2466.48 is associated with stabilizing the furan ring common in many adenosine A2 a antagonists. The binding pocket also includes four ordered water molecules.

The extracellular regions of the A2a receptor differ markedly from the Preceptors. The second extracellular loop does not form an a-helix but rather a random coil constrained by three disulfide bonds. A further disulfide bond links the third extracellular loop with the top of TM6, producing a kinked structure. This array of disulfide bonds helps to keep an open entrance to the ligand binding pocket, presumably allowing access of the natural ligand from the extracellular space.

As with the PAR structures, the ionic lock is also absent in the A2a structure. The absence of the lock in three structures starts to bring into question the relevance of the ionic lock as a mechanism for stabilizing the inactive state of GPCRs, other than rhodopsin. Instead, the adenosine A2a receptor and p1AR contain a network of hydrogen bond interactions between Asp101349 and TM2, and the second ICL. The role of Arg102350 in this situation may be to stabilize the deprotonated Asp101349.

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