Purification, functional characterization, and crystallization of the ligand binding domain of the retinoid X receptor

Protein Expr Purif. 1995 Oct;6(5):604-8. doi: 10.1006/prep.1995.1079.

Abstract

The ligand binding domain (LBD) of the human retinoid X receptor alpha (hRXR alpha) was overproduced in Escherichia coli and purified to more than 95% purity and functional homogeneity. Circular dichroism spectra of the purified RXR alpha LBD indicated that the protein was composed predominantly of alpha-helical structures and coils. Crystals were grown from ammonium citrate using the vapor diffusion method against a reservoir containing 100 mM Pipes (pH 7.0) and 1.5 M ammonium citrate. They belong to the hexagonal space group P6(3)22 with unit cell parameters a = b = 110.8 A and c = 109.9 A, alpha = beta = 90 degrees, gamma = 120 degrees, and they diffract X rays to a resolution limit of 2.5 A using synchrotron radiation. The asymmetric unit of the crystals contains one molecule with a solvent content of approximately 55% and a Vm value of 3.6 A3/dalton.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Base Sequence
  • Binding Sites
  • Chromatography, Gel
  • Chromatography, Ion Exchange
  • Circular Dichroism
  • Cloning, Molecular
  • Crystallization
  • Crystallography, X-Ray*
  • Escherichia coli / genetics
  • Escherichia coli / metabolism
  • Fluorescence
  • Histidine / chemistry
  • Histidine / metabolism
  • Humans
  • Ligands
  • Magnetic Resonance Spectroscopy
  • Molecular Sequence Data
  • Receptors, Retinoic Acid / chemistry*
  • Receptors, Retinoic Acid / isolation & purification
  • Receptors, Retinoic Acid / metabolism
  • Retinoid X Receptors
  • Transcription Factors / chemistry*
  • Transcription Factors / isolation & purification
  • Transcription Factors / metabolism

Substances

  • Ligands
  • Receptors, Retinoic Acid
  • Retinoid X Receptors
  • Transcription Factors
  • Histidine