The multitude of G-protein coupled receptor (GPR) superfamily cDNAs recently isolated has exceeded the number of receptor subtypes anticipated by pharmacological studies. Analysis of the sequence similarities and unique features of the members of this family is valuable for designing strategies to isolate related cDNAs, for developing hypotheses concerning substrate-ligand and receptor-effector interactions, and for understanding the evolution of these genes. We have compiled and aligned the 74 unique amino acid sequences published to date and review the present understanding of the structural motifs contributing to ligand binding and G-protein coupling.

Sequence alignment of the G-protein coupled receptor superfamily.

Dopamine receptors: Molecular biology, biochemistry and behavioural aspects

Modelling G-protein-coupled receptors for drug design.

The introduction of clozapine has given clinicians a unique agent for treating patients with schizophrenia that is refractory to other neuroleptics. Despite its efficacy, the drug continues to be prescribed with trepidation due to the incidence of agranulocytosis. This article reviews the pharmacokinetic and pharmacological properties of clozapine and the clinical implications for monitoring plasma concentrations. Various assays have been developed for clozapine that include gas-liquid chromatography, radioimmunoassay and high performance liquid chromatography. Only a few studies have examined the pharmacokinetics of clozapine in patients with schizophrenia. These studies have revealed a wide interpatient variability in pharmacokinetic parameters that include: time to reach peak plasma concentrations 1.1 to 3.6h; elimination half-life 9.1 to 17.4h; clearance 8.7 to 53.3 L/h; and a volume of distribution of 1.6 to 7.3 L/kg. Clozapine is metabolised via the hepatic microsomal enzyme system into 2 principle metabolites: demethyl-clozapine and clozapine N-oxide. Urine samples have reported the ratio of clozapine:demethyl:N-oxide to be 1:1:2. The clozapine N-oxide binding affinity with 3H-haloperidol was 4 times lower than clozapine and its conversion back to clozapine is hypothesised. Although the exact pharmacological mechanism of action of clozapine is not fully understood, the drug does possess significant binding affinity for different dopamine receptors, with recent evidence supporting binding to the D4 receptor subtype. Clozapine transiently increases serum prolactin levels with minimal changes in homovanillic acid plasma levels. Limited studies investigating the relationship between clinical response and plasma clozapine concentrations have investigated the range between 100 and 800 micrograms/L. In the treatment of patients with refractory schizophrenia, a minimum concentration of 350 micrograms/L was suggested as needed. The occurrence of agranulocytosis could have a genetic basis and patients should be rigorously monitored during treatment. The incidence of tardive dyskinesia and extrapyramidal side effects is minimal. Clozapine can lower the seizure threshold in a dose- and time-dependent manner. Careful patient selection and monitoring are required when clozapine therapy is used in patients with schizophrenia.

Pharmacokinetics and Pharmacodynamics of Clozapine

Computational modeling approaches to structure-function analysis of G protein-coupled receptors.

A three-dimensional model of the dopamine D2 receptor, assumed to be a target of antipsychotic drug action, was constructed from its amino acid sequence. The model was based on structural similarities within the super-family of guanine nucleotide-binding regulatory (G) protein-coupled neuroreceptors and has seven alpha-helical transmembrane segments that form a central core with a putative ligand-binding site. The space between two residues postulated to be involved in agonist binding, Asp-80 and Asn-390, perfectly accommodated an anti-dopamine molecule. Molecular electrostatic potentials were mainly negative on the synaptic side of the receptor model and around aspartate residues lining the central core and positive in the cytoplasmic domains. The docking of dopamine into a postulated binding site was examined by molecular dynamics simulation. The protonated amino group became oriented toward negatively charged aspartate residues in helix 2 and helix 3, whereas the dopamine molecule fluctuated rapidly between different anti and gauche conformations during the simulation. The receptor model suggests that protonated ligands are attracted to the binding site by electrostatic forces and that protonated agonists may induce conformational changes in the receptor, leading to G-protein activation, by increasing the electrostatic potentials near Asp-80.

https://www.pnas.org/content/pnas/88/18/8111.full.pdf

Ingebright Sylte

Papers

Molecular dynamics of dopamine at the D2 receptor.