Expression, purification, and characterization of multiple, multifunctional human glucocorticoid receptor proteins.
The glucocorticoid receptor (GR) is a nuclear receptor protein that plays a central role in glucose homeostasis, the stress response, control of the hypothalamic-pituitary-adrenal axis, and immuno-inflammatory processes via binding of the natural steroid, cortisol. GR is a well-validated drug target and continues to be an important target for new drug discovery efforts. Here, we describe a basic and simple method for Escherichia coli expression and purification of a variety of human GR proteins that contain all three of the functional domains of the protein: the activation function-1 domain, the DNA-binding domain, and the ligand-binding domain. We present characterization data to show that these purified, multifunctional GR proteins are active for ligand, coactivator, and DNA-binding. The work presented here should serve as a reference for future mechanistic, structural and drug discovery efforts that require purified, full or near full length, GR protein.
Simmons
Catherine A
CA
Molecular Discovery Research, GlaxoSmithKline, Research Triangle Park, NC 27709, USA.
Bledsoe
Randy K
RK
Guex
Nicolas
N
Pearce
Kenneth H
KH
D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D006801:Humans; D008024:Ligands; D011965:Receptors, Glucocorticoid / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification*
D008024:Ligands; D011965:Receptors, Glucocorticoid