Kinetic analysis of estrogen receptor homo- and heterodimerization in vitro

J Steroid Biochem Mol Biol. 2003 Feb;84(2-3):141-8. doi: 10.1016/s0960-0760(03)00023-2.

Abstract

The coexistence of ERalpha and ERbeta suggests that active receptor complexes are present as homo- or heterodimers. In addition each of three forms of active receptors may trigger different cellular responses. A real-time biosensor based on surface plasmon resonance was used as instrument to determine binding kinetics of homo- and heterodimerization of estrogen receptor alpha and beta. Partially purified full-length estrogen receptor alpha was expressed intracellularly as a C-terminal fusion to a hexa-histidine tag using the baculovirus-expression system. Purified estrogen receptor alpha and beta without tags were used as partners in the dimerization process. An association rate constant of 3.6 x 10(3) to 1.5 x 10(4)M(-1)s(-1) for the homodimer formation of ERalpha and 5.7 x 10(3) to 1.5 x 10(4)M(-1)s(-1) for the heterodimer formation was found assuming a pseudo first-order reaction kinetic. The equilibrium dissociation constant for homodimerization of ERalpha was 2.2 x 10(-8) to 5.4 x 10(-8) and 1.8 x 10(-8) to 2.6 x 10(-8)M for the heterodimer formation. The homo- and heterodimer formation was characterized by a slow association kinetics and kinetic rate constants were within the same range.

MeSH terms

  • Animals
  • Biosensing Techniques
  • Cell Line
  • Dimerization
  • Estradiol / metabolism
  • Estrogen Receptor alpha
  • Humans
  • Insecta
  • Kinetics
  • Ligands
  • Plasmids / metabolism
  • Protein Binding
  • Protein Structure, Tertiary
  • Receptors, Estrogen / chemistry*
  • Receptors, Estrogen / metabolism
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / metabolism
  • Time Factors
  • Transfection

Substances

  • Estrogen Receptor alpha
  • Ligands
  • Receptors, Estrogen
  • Recombinant Proteins
  • Estradiol