Solubilization and characterization of PGE2 receptor in porcine ciliary epithelium.
PGE2 binding sites or receptors of porcine ciliary nonpigmented epithelial (NPE) and pigmented epithelial (PE) membranes were solubilized with detergents (CHAPS and Triton X100). From the Scatchard plots of PGE2 binding to CHAPS-solubilized proteins, the Kd and Bmax values were calculated to be 35 nM and 470 fmol/mg protein for NPE protein and 65 nM and 430 fmol/mg protein for PE protein, respectively. On the basis of the Kd and Bmax values, the solubilized receptor proteins correspond to PGE2 binding sites of the membranes which have previously been shown to be coupled to adenylate cyclase inhibition. For both NPE and PE proteins, the order of binding potency was PGE2 > PGF2 alpha > PGD2. By gel filtration chromatography of NPE and PE proteins, the molecular mass of the major PGE2 binding peak was estimated to be about 150 KDa when solubilized in CHAPS and 46 KDa for Triton X100 extracts. The Bmax values of membrane-associated binding proteins were increased by GTP, indicating a close association of the PGE2 binding sites with a GTP-binding protein. However, GTP did not affect the Bmax values of detergent-solubilized receptor proteins.
Sano
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Kresge Eye Institute, Department of Ophthalmology, Wayne State University School of Medicine, Detroit, Michigan.
Shichi
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H
D000818:Animals; D001667:Binding, Competitive / Q000187:drug effects; D002793:Cholic Acids; D002924:Ciliary Body / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D015232:Dinoprostone / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D008565:Membrane Proteins / Q000187:drug effects / Q000302:isolation & purification / Q000378:metabolism; D017830:Octoxynol; D010857:Pigment Epithelium of Eye / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D018078:Receptors, Prostaglandin E / Q000302:isolation & purification / Q000378:metabolism*; D013552:Swine
D002793:Cholic Acids; D008565:Membrane Proteins; D018078:Receptors, Prostaglandin E; D006160:Guanosine Triphosphate; D017830:Octoxynol; D015232:Dinoprostone; C028213:3-((3-cholamidopropyl)dimethylammonium)-1-propanesulfonate