Histidine 6.55 is a major determinant of ligand-biased signaling in dopamine D2L receptor

Mol Pharmacol. 2011 Mar;79(3):575-85. doi: 10.1124/mol.110.068106. Epub 2010 Dec 16.

Abstract

In our previous studies, we demonstrated that the mutation of His393(6.55) to alanine results in an increased affinity of 1,4-disubstituted phenylpiperazines to the dopamine D(2L) receptor. This change most likely accounts for the reduced steric hindrance in this part of the binding pocket. In this work, we investigated the role of the steric hindrance imposed by the residue His393(6.55) for the receptor activation modulated by 1,4-disubstituted aromatic piperidines/piperazines. Site-directed mutagenesis and ligand modifications were used to probe the structural basis of ligand efficacy. The operational model of agonism was used to quantify the ligand bias between the ability of compounds to inhibit cAMP accumulation and stimulate extracellular signal-regulated kinase 1/2 (ERK1/2) phosphorylation. Whereas substantial ligand-biased signaling was observed for the D(2L) wild-type receptor, an overall increase in agonism was observed for the D(2L) H393(6.55)A mutant without noteworthy functional selectivity. Targeted chemical modification of the phenylpiperazine moiety at the site of its interaction with the residue His393(6.55) led to the functionally selective ligand {3-[4-(2,3-dihydro-benzofuran-7-yl)-piperazin-1-yl]-propyl}-pyrazol[1,5-a]pyridine-3-carboxamide (FAUC350) that has distinct signaling profiles toward adenylyl cyclase and ERK1/2. FAUC350 behaves as an antagonist in the inhibition of cAMP accumulation and as a partial agonist in the stimulation of ERK1/2 phosphorylation (efficacy = 55%). Overall, the residue His393(6.55) and proximate molecular substructures of receptor ligands were identified to be crucial for multidimensional ligand efficacy.

MeSH terms

  • Adenylyl Cyclases / drug effects
  • Adenylyl Cyclases / metabolism
  • Animals
  • CHO Cells
  • Catalytic Domain / drug effects
  • Catalytic Domain / physiology
  • Cricetinae
  • Cricetulus
  • Cyclic AMP / metabolism
  • Cyclic AMP / physiology
  • Dopamine Agonists / pharmacology
  • Dopamine Antagonists / pharmacology
  • Enzyme-Linked Immunosorbent Assay
  • Extracellular Signal-Regulated MAP Kinases / drug effects
  • Extracellular Signal-Regulated MAP Kinases / metabolism
  • Extracellular Signal-Regulated MAP Kinases / physiology
  • Histidine
  • Ligands
  • Mutagenesis, Site-Directed
  • Phosphorylation
  • Radioligand Assay
  • Receptors, Dopamine D2 / chemistry
  • Receptors, Dopamine D2 / drug effects
  • Receptors, Dopamine D2 / metabolism
  • Receptors, Dopamine D2 / physiology*
  • Signal Transduction* / drug effects
  • Signal Transduction* / physiology

Substances

  • Dopamine Agonists
  • Dopamine Antagonists
  • Ligands
  • Receptors, Dopamine D2
  • dopamine D2L receptor
  • Histidine
  • Cyclic AMP
  • Extracellular Signal-Regulated MAP Kinases
  • Adenylyl Cyclases