Large-scale purification and characterization of human parathyroid hormone-1 receptor stably expressed in HEK293S GnTI − cells

Abstract

Human parathyroid hormone-1 receptor (hPTHR1) belongs to class II of the G protein-coupled receptor (GPCR) family, whose members all contain a seven-transmembrane helix domain. The receptor regulates bone metabolism through interactions with its ligand, human parathyroid hormone (hPTH). For structural studies of the hPTHR1/hPTH complex, we constructed a mammalian cell line to stably express recombinant hPTHR1 in large-scale. The receptor was solubilized with dodecyl maltoside and purified with affinity chromatography. The purified receptor displayed restricted N-glycosylation as expected. Functionality was demonstrated: the hPTHR1 retained affinity for bPTH-(1–34) and specifically cross-linked to a radioiodinated bPTH-(1–34) analog. This work describes an approach for preparing milligram-scale quantities of receptor for elucidation of the structural biology of this seven-transmembrane GPCR.