Structure of the human histamine H1 receptor complex with doxepin

Abstract

The biogenic amine histamine is an important pharmacological mediator involved in pathophysiological processes such as allergies and inflammations. Histamine H(1) receptor (H(1)R) antagonists are very effective drugs alleviating the symptoms of allergic reactions. Here we show the crystal structure of the H(1)R complex with doxepin, a first-generation H(1)R antagonist. Doxepin sits deep in the ligand-binding pocket and directly interacts with Trp 428(6.48), a highly conserved key residue in G-protein-coupled-receptor activation. This well-conserved pocket with mostly hydrophobic nature contributes to the low selectivity of the first-generation compounds. The pocket is associated with an anion-binding region occupied by a phosphate ion. Docking of various second-generation H(1)R antagonists reveals that the unique carboxyl group present in this class of compounds interacts with Lys 191(5.39) and/or Lys 179(ECL2), both of which form part of the anion-binding region. This region is not conserved in other aminergic receptors, demonstrating how minor differences in receptors lead to pronounced selectivity differences with small molecules. Our study sheds light on the molecular basis of H(1)R antagonist specificity against H(1)R.

Fig. 1. Structure of H₁R complex with doxepin

(a) Ribbon representation of the H₁R structure. Doxepin is shown as yellow spheres whereas the phosphate ion as spheres with carbon, and oxygen atoms colored orange and red, respectively. Disulfide bonds are shown as yellow sticks, and Trp428 and Asp107 as pink sticks. Three conserved motifs D(E)R^3.50Y, CWxP^6.50 and NP^7.50xxY are highlighted in blue. (b) Superimposition of the H₁R (green) and β₂AR (cyan) structures. (c) Same as (b) but with the D3R structure colored magenta.

Fig. 2. Comparison of the structures of H₁R, β₂AR and D3R

(a) Proline-induced kink in helix IV (H₁R: green, β₂AR: cyan, D3R: magenta). The side chain of Trp158^4.56 and Pro161^4.59 of H₁R and the equivalent residues of β₂AR (Ser165^4.57 and Pro168^4.60) and of D3R (Ser165^4.57 and Pro167^4.59) are also shown. (b) Variations in the D(E)RY motif structures of H₁R, β₂AR and D3R colored in green, cyan and magenta, respectively. Side chains of Asp124^3.49, Arg125^3.50, Glu410^6.30 of H₁R and the equivalents of β₂AR and D3R are represented as stick models. For H₁R, Gln416^6.36, which forms a hydrogen bond with Arg125^3.50 , are also shown. Possible hydrogen bonds are indicated by dotted lines.

Fig. 3. Binding interactions of doxepin

(a) Doxepin is shown as sticks with yellow carbon atoms, whereas the contact residues within 4 Å are shown with grey carbon atoms. Nitrogen and oxygen atoms are colored blue and red, respectively. Hydrogen bonds/salt bridges are indicated as blue dotted lines. (b) Doxepin binding interactions. Hydrophobic interactions are shown in black dotted lines. (c) Ligand binding positions in non-rhodopsin GPCRs. Carbon atoms of doxepin (H₁R) are shown in yellow, carazolol (β₂AR) in cyan, eticlopride (D3R) in magenta and ZM241385 (A_2AAR) in grey. (d) Structure of the anion-binding region with a phosphate ion.

Fig.4. Interactions of second-generation selective H₁R-antagonists with the H₁R ligand-binding pocket

Conformation of each complex was predicted by global optimization of the ligand in the all-atom flexible H₁R model ^,, based on the H₁R-doxepin complex structure. Carbon atoms for (a) Z-olopatadine co-bound with the phosphate ion, (b) acrivastine, (c) R-cetirizine (levocetirizine), and (d) fexofenadine are colored magenta. Nitrogen and oxygen atoms are colored blue and red, respectively. Ligand contact residues of H₁R are shown with grey carbon atoms; parts of helices III, IV and ECL2 are not displayed for clarity. Hydrogen bonds are shown in cyan.