Fig. 5.

Purification of inner mitochondrial membrane AT₂Rs. (A) Whole-liver homogenate fractionations up to the inner mitochondrial membrane were subjected to 12% SDS/PAGE and immunoblotting with anti-AT₂R as well as anti-Na⁺/K⁺ ATPase, anti-VDAC, and anti-ATP synthase β for detecting cell membrane, outer mitochondrial membrane, and inner mitochondrial membrane markers, respectively. AT₂Rs tracked with inner mitochondrial membrane marker ATP synthase β, consistent with inner mitochondrial membrane localization of AT₂Rs. (B) Integrated densitometric band analysis of immunoblots demonstrating fold enrichment of AT₂Rs, inner mitochondrial membrane marker (ATP synthase), outer mitochondrial membrane marker (VDAC), and plasma membrane marker (Na/K ATPase) with mitochondrial purification. (C) Percentage enrichment of AT₂Rs with cellular subfractions through mitochondrial purification. Fractions: 1, whole-cell lysate; 2, postnuclear (480 × g); 3, postdifferential centrifugation (7,700 × g); 4, post-HistoDenz gradient centrifugation (50,500 × g); 5, postsucrose gradient centrifugation (77,000 × g).