A nanomolar-potency small molecule inhibitor of regulator of G-protein signaling proteins

Biochemistry. 2011 Apr 19;50(15):3181-92. doi: 10.1021/bi1019622. Epub 2011 Mar 29.


Regulators of G-protein signaling (RGS) proteins are potent negative modulators of signal transduction through G-protein-coupled receptors. They function by binding to activated (GTP-bound) Gα subunits and accelerating the rate of GTP hydrolysis. Modulation of RGS activity by small molecules is an attractive mechanism for fine-tuning GPCR signaling for therapeutic and research purposes. Here we describe the pharmacologic properties and mechanism of action of CCG-50014, the most potent small molecule RGS inhibitor to date. It has an IC(50) for RGS4 of 30 nM and is >20-fold selective for RGS4 over other RGS proteins. CCG-50014 binds covalently to the RGS, forming an adduct on two cysteine residues located in an allosteric regulatory site. It is not a general cysteine alkylator as it does not inhibit activity of the cysteine protease papain at concentrations >3000-fold higher than those required to inhibit RGS4 function. It is also >1000-fold more potent as an RGS4 inhibitor than are the cysteine alkylators N-ethylmaleimide and iodoacetamide. Analysis of the cysteine reactivity of the compound shows that compound binding to Cys(107) in RGS8 inhibits Gα binding in a manner that can be reversed by cleavage of the compound-RGS disulfide bond. If the compound reacts with Cys(160) in RGS8, the adduct induces RGS denaturation, and activity cannot be restored by removal of the compound. The high potency and good selectivity of CCG-50014 make it a useful tool for studying the functional roles of RGS4.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Alkylation / drug effects
  • Biocatalysis
  • Cell Survival
  • Cysteine / metabolism
  • Flow Cytometry
  • GTP-Binding Protein alpha Subunits, Gi-Go / metabolism
  • HEK293 Cells
  • Humans
  • Models, Molecular
  • Protein Conformation
  • RGS Proteins / antagonists & inhibitors*
  • RGS Proteins / chemistry
  • RGS Proteins / metabolism
  • Substrate Specificity
  • Thiadiazoles / chemistry*
  • Thiadiazoles / metabolism
  • Thiadiazoles / pharmacology*
  • Thiazolidinediones / chemistry*
  • Thiazolidinediones / metabolism
  • Thiazolidinediones / pharmacology*


  • 4-(4-fluorobenzyl)-2-p-tolyl-1,2,4-thiadiazolidine-3,5-dione
  • RGS Proteins
  • RGS8 protein, human
  • Thiadiazoles
  • Thiazolidinediones
  • RGS4 protein
  • GTP-Binding Protein alpha Subunits, Gi-Go
  • Cysteine