Overexpression and Functional Stabilization of Recombinant Human Lysophosphatidic Acid Receptor 1 Using an Amphiphatic Polymer

Human lysophosphatidic acid receptor 1 (LPA(1)) is a G-protein coupled receptor that mediates various biological functions such as proliferation, platelet aggregation, smooth muscle contraction, and tumor cell invasion. For dissection of the molecular function of LPA(1), a recombinant LPA(1) was overexpressed in Escherichia coli membrane fractions and purified to homogeneity by single affinity chromatography. The purified LPA(1) was stabilized with an amphiphilic polymer that was synthesized by the coupling of octylamine, glucosamine, and diethylaminoproylamine at the carboxylic groups of poly-gamma-glutamic acid. The complex of purified LPA(1) and amphiphilic polymer showed a monodisperse oligomer and specific binding to LPA with apparent K-i values of 30 mu M. Compared with the G(s) protein, it also showed selective binding to the alpha subunit of the G(i) protein. These results indicate that recombinant LPA(1) in an amphiphilic polymer complex has an active conformation for interaction with ligands and G-proteins.