Fig. 3.

DDM solubilises considerable amounts of inactive AT₁R produced in the baculovirus expression system. (a) Western blot of DDM-solubilised AT₁R, with equal amounts of active receptor per sample (lanes 2, 3 and 5–10). The blot was probed with an anti-pentaHis-HRP conjugated antibody. Lane 1, iHEK parental cells; lanes 2 and 3, iHEK(AT₁R-GFP-H₁₀) stable clonal cell line; lane 4, uninfected Sf9 cells; lanes 5–10, bvAT₁R-H₁₀ infected insect cells. N-Linked glycosylation was removed using PNGase F where indicated (+). AT₁R was expressed either in the stable mammalian cell line iHEK(AT₁R-GFP-H₁₀) or by using the recombinant baculoviruses bvAT₁R-H₁₀ and bvAT₁R-LS-H₁₀ to infect Sf9, Sf21 and Hi5 cells as indicated. iHEK cell lines were induced with 1 μg/ml tetracycline for 24 h and insect cells were infected with recombinant baculovirus for 48 h. The amount of functional AT₁R was determined by measuring specific binding of the antagonist [¹²⁵I]Sar¹.