Expression of the Ligand-Binding Domain-Containing Region of Retinoic Acid Receptors α, β and γ in Escherichia coli and Evaluation of Ligand-Binding Selectivity

Abstract

The complete molecule or the ligand-binding domain-containing region of each of the three subtypes of human retinoic acid receptors (hRARα, hRARβ and hRARγ) was expressed in Escherichia coli. The expressed recombinant RARs (rRARs: rRARα/E, rRARβ/E and rRARγ) showed nearly the same magnitude of binding affinity toward [3H]retinoic acid (RA) as hRARs extracted from human cells (Ka values: 6.0×109 M-1 for rRARα/E and 2.7×1010 M-1 for both rRARβ/E and rRARγ). Therefore, the ligand-binding selectivity of the rRARs toward RA and synthetic retinoids (4-(5, 6, 7, 8-tetrahydro-5, 5, 8, 8-tetrametyl-2-naphthalenylcarbamoyl)benzoic acid (Am80), (E)-4-[3-(3, 5-di-tert-butylphenyl)-3-oxo-1-propenyl]benzoic acid (Ch55)) was examined. Am80 bound rRARα/E preferentially and showed no binding activity toward rRARγ, which is consistent with the case of hRARγ. Ch55 bound all three subtypes of rRARs, preferentially rRARβ/E. These results suggest that the intrinsic nature of the binding of each retinoid can be investigated by usage of the rRARs. However, rRARs show quantitatively different ligand-selectivity from that of hRARs: RA showed higher binding affinity toward rRARs than both Am80 and Ch55, but Ch55 binds all three subtypes of hRARs stronger than RA and Am80, which binds hRARβ stronger than RA. © 1993, The Pharmaceutical Society of Japan. All rights reserved.