PROSTAGLANDIN-E1 RECEPTOR FROM MOUSE MASTOCYTOMA P-815 CELLS COUPLES TO 60-KDA GTP-BINDING PROTEIN

The stable [H-3]prostaglandin E1 (PGE1)-bound receptor, which couples to 60 kDa GTP-binding protein, from membranes of mouse mastocytoma P-815 cells has been purified and characterized. When the membranes were preincubated with [H-3]PGE1 for 60 min at 37-degrees-C, the dissociation of the ligand from the receptor was remarkably decreased, even in the presence of GTP-gamma-S. The stable [H-3]PGE1-bound receptor complex was solubilized with 6% digitonin. The solubilized [H-3]PGE1 receptor was eluted with [S-35]GTP-gamma-S binding activity from an Ultrogel AcA44 column. The fractions containing activities of both [H-3]PGE1 and [S-35]GTP-gamma-S bindings were further purified by column chromatographies on wheat germ agglutinin (WGA)-agarose and phenyl-Sepharose CL-4B. The partially purified [H-3]PGE1-bound receptor was affinity-labeled with [C-14]5'-p-fluorosulfonylbenzoylguanosine and a protein with a molecular mass of 60 kDa was detected. These results suggest that the ligand-bound PGE1 receptor of P-815 cells associates with a novel GTP-binding protein with a molecular mass of 60 kDa.