Partial purification and biochemical characterization of a membrane glucocorticoid receptor from an amphibian brain

A membrane receptor for corticosterone (mGR) in the brain of the roughskin newt (Taricha granulosa) has been previously identified. This manuscript re ports the evaluation of several chromatographic resins for enrichment of th e newt mGR solubilized from neuronal membranes. A protein with an apparent molecular weight of 63 kDa was purified to near homogeneity following seque ntial purification using ammonium sulfate fractionation, wheat germ aggluti nin (WGA)-agarose chromatography, hydroxylapatite chromatography, and an im mobilized ligand affinity resin (Corticosterone-Sepharose). Other studies e mployed a novel protein differential display strategy and a photoaffinity l abeling strategy to visualize candidate receptor proteins following SDS-PAG E. Both of these techniques also identified a 63 kDa protein, agreeing with the estimation of molecular weight from the purification data. Furthermore , the use of 2D SDS-PAGE following the photolabeling procedure showed the c andidate 63 kDa protein to have a pi of approximately 5.0. Taken together t hese data suggest that the newt mGR is an acidic glycoprotein with an appar ent molecular weight of 63 kDa. Because these characteristics of newt mGR a re inconsistent with the characteristics of intracellular glucocorticoid re ceptors, these two receptor proteins are apparently distinct. (C) 2000 Else vier Science Ltd. All rights reserved.