Figure: The beta 2 AR (green) bound to adrenaline (orange) and stabilized in an active state by an engineered nanobody (blue).
The groups of Brian Kobilka, Bill Weis, and Chris Garcia at Stanford University determined the adrenaline-activated structure of the beta 2 adrenoceptor stabilized with an engineered nanobody. The beta 2 adrenergic receptor (beta 2 AR) is a G-protein-coupled receptor (GPCR) that mediates cellular responses to adrenaline. Kobilka, Weis, and collaborators have previously obtained structures of the beta 2 AR in inactive and active states, but it had not been possible to capture an active-state of a GPCR bound to its native neurotransmitter. Crystal structures of agonist-bound GPCRs have relied on the use of either exceptionally high-affinity agonists or receptor stabilization by mutagenesis. Many natural agonists such as adrenaline bind with relatively low affinity, and they are often chemically unstable. Using directed evolution, they engineered a high-affinity camelid antibody fragment (nanobody) that stabilizes the active state of the beta 2 AR, and used this to obtain crystal structures of the activated receptor bound to three chemically distinct agonists: the ultrahigh-affinity agonist BI167107, the high-affinity catecholamine agonist hydroxybenzyl isoproterenol, and the low-affinity endogenous agonist adrenaline. The crystal structures reveal a highly conserved overall ligand recognition and activation mode despite diverse ligand chemical structures and affinities that range from 100 nM to ~80 pM. Overall, the adrenaline-bound receptor structure is similar to the others, but it has substantial rearrangements in extracellular loop three and the extracellular tip of transmembrane helix 6. These structures also reveal a water-mediated hydrogen bond between two conserved tyrosines that appears to stabilize the active state of the beta 2 AR and related GPCRs.
Citation: Ring, AM, Manglik, A, Kruse, AC, Enos, MD, Weis, WI, Garcia, KC, Kobilka, BK. Adrenaline-activated structure of [beta subscript 2]-adrenoceptor stabilized by an engineered nanobody, Nature 502, 575-579 (2013). DOI: 10.1038/nature12572