Investigating Allosteric Coupling Mechanism in Constitutively Active Human Glucocorticoid Receptor Two Domain Construct

Abstract

Glucocorticoid receptor (GR) is a hormone dependent nuclear transcription factor. It plays key roles in organ development, metabolite homeostasis, and stress and inflammatory response. GR is composed of three major domains, the intrinsically disordered (ID) N-terminal domain (NTD), the DNA binding domain (DBD) and the ligand binding domain (LBD). Several human GR translational isoforms differing only in the lengths of their ID NTDs with finely tunable transcription activity have been identified.In my project, through thermodynamic analysis of the ID NTD constructs of different translational isoforms, we found that full length GR NTD can be viewed as having at least two thermodynamically coupled regions, a functional (F) region, indispensable for GR transcriptional activity, and a regulatory (R) region, whose length serves to regulate the stability of NTD, and thus the activity of GR. F region and R region are negatively coupled to each other [1].Truncation of LBD from the full length receptor removes the hormone dependence of GR transcription activity, and generates the constitutively active GR NTD-DBD two domain constructs. From investigating the NTD-DBD two domain constructs of different translational isoforms by in vitro biophysics assays and in vivo luciferase activity assay, we are on the way figuring out the coupling mechanism between the DBD and ID NTD.Allosteric coupling mechanisms derived from our experimental data makes it possible to build the Ensemble Allosteric Coupling Model (EAM) to explain the finely tunable activity of different GR translational isoforms, which cannot be accounted for by the binding energetics between the different isoforms and the GR response element.[1] Jing Li, et al. Thermodynamic dissection of the intrinsically disordered N-terminal domain of human glucocorticoid receptor. J Biol Chem. 2012.