The angiotensin II receptors AT₁R and AT₂R serve as key components of the renin–angiotensin–aldosterone system. AT₁R has a central role in the regulation of blood pressure, but the function of AT₂R is unclear and it has a variety of reported effects. To identify the mechanisms that underlie the differences in function and ligand selectivity between these receptors, here we report crystal structures of human AT₂R bound to an AT₂R-selective ligand and to an AT₁R/AT₂R dual ligand, capturing the receptor in an active-like conformation. Unexpectedly, helix VIII was found in a non-canonical position, stabilizing the active-like state, but at the same time preventing the recruitment of G proteins or β-arrestins, in agreement with the lack of signalling responses in standard cellular assays. Structure–activity relationship, docking and mutagenesis studies revealed the crucial interactions for ligand binding and selectivity. Our results thus provide insights into the structural basis of the distinct functions of the angiotensin receptors, and may guide the design of new selective ligands.

Angiotensin II type 1 receptor (AT₁R) is the primary blood pressure regulator. AT₁R blockers (ARBs) have been widely used in clinical settings as anti-hypertensive drugs and share a similar chemical scaffold, although even minor variations can lead to distinct therapeutic efficacies toward cardiovascular etiologies. The structural basis for AT₁R modulation by different peptide and non-peptide ligands has remained elusive. Here, we report the crystal structure of the human AT₁R in complex with an inverse agonist olmesartan (Benicar^TM), a highly potent anti-hypertensive drug. Olmesartan is anchored to the receptor primarily by the residues Tyr-35^1.39, Trp-84^2.60, and Arg-167^ECL2, similar to the antagonist ZD7155, corroborating a common binding mode of different ARBs. Using docking simulations and site-directed mutagenesis, we identified specific interactions between AT₁R and different ARBs, including olmesartan derivatives with inverse agonist, neutral antagonist, or agonist activities. We further observed that the mutation N111^3.35A in the putative sodium-binding site affects binding of the endogenous peptide agonist angiotensin II but not the β-arrestin-biased peptide TRV120027.

Serial femtosecond crystallography of the human δ-opioid receptor in complex with an endomorphin-derived peptide reveals interactions that are important for understanding the pharmacology of opioid peptides and developing analgesics with reduced side effects. Bifunctional μ- and δ-opioid receptor (OR) ligands are potential therapeutic alternatives, with diminished side effects, to alkaloid opiate analgesics. We solved the structure of human δ-OR bound to the bifunctional δ-OR antagonist and μ-OR agonist tetrapeptide H-Dmt-Tic-Phe-Phe-NH2 (DIPP-NH2) by serial femtosecond crystallography, revealing a cis-peptide bond between H-Dmt and Tic. The observed receptor-peptide interactions are critical for understanding of the pharmacological profiles of opioid peptides and for development of improved analgesics.

β-N-Acetyl-d-hexosaminidase has been postulated to have a specialized function. However, the structural basis of this specialization is not yet established. OfHex1, the enzyme from the Asian corn borer Ostrinia furnacalis (one of the most destructive pests) has previously been reported to function merely in chitin degradation. Here the vital role of OfHex1 during the pupation of O. furnacalis was revealed by RNA interference, and the crystal structures of OfHex1 and OfHex1 complexed with TMG-chitotriomycin were determined at 2.1 Å. The mechanism of selective inhibition by TMG-chitotriomycin was related to the existence of the +1 subsite at the active pocket of OfHex1 and a key residue, Trp⁴⁹⁰, at this site. Mutation of Trp⁴⁹⁰ to Ala led to a 2,277-fold decrease in sensitivity toward TMG-chitotriomycin as well as an 18-fold decrease in binding affinity for the substrate (GlcNAc)₂. Although the overall topology of the catalytic domain of OfHex1 shows a high similarity with the human and bacterial enzymes, OfHex1 is distinguished from these enzymes by large conformational changes linked to an “open-close” mechanism at the entrance of the active site, which is characterized by the “lid” residue, Trp⁴⁴⁸. Mutation of Trp⁴⁴⁸ to Ala or Phe resulted in a more than 1,000-fold loss in enzyme activity, due mainly to the effect on k_cat. The current work has increased our understanding of the structure-function relationship of OfHex1, shedding light on the structural basis that accounts for the specialized function of β-N-acetyl-d-hexosaminidase as well as making the development of species-specific pesticides a likely reality.