title: Biochemical characterization of a 22-kDa high affinity antiischemic drug-binding polypeptide in the endoplasmic reticulum of guinea pig liver: potential common target for antiischemic drug action.
abstract: The phenylalkylamine emopamil prevents brain damage due to experimental cerebral ischemia. Stereoselective, high affinity, binding sites for (-)-[3H]emopamil in guinea pig brain cortex and liver membranes have been proposed to mediate its antiischemic effect. Using [N-methyl-3H]LU49888 as a photoaffinity probe we now provide evidence that the cation-sensitive emopamil binding site is localized on a 22-kDa polypeptide in guinea pig liver, kidney, lung, and adrenal gland. This 22-kDa polypeptide binds other antiischemic drugs with high affinity and is a nonglycosylated integral membrane protein of the endoplasmic reticulum. It can be solubilized with digitonin without changes in its drug-binding properties. The solubilized binding activity has a sedimentation coefficient of 12.0 +/- 0.4 S and an apparent Stokes radius of 6.0 +/- 0.1 nm. From these data it is concluded that the 22-kDa polypeptide is associated in a larger oligomeric complex with a molecular mass of at least 84 kDa. [N-methyl-3H]LU49888 also specifically labels a second 27-kDa polypeptide in the endoplasmic reticulum, which can be distinguished from the 22-kDa polypeptide by its pharmacological and hydrodynamic properties. The photolabeled 22-kDa polypeptide was partially purified under denaturating conditions. This will allow the further structural analysis of this putative target for antiischemic drugs.
authors.affiliation: Institut für Biochemische Pharmakologie, Universität Innsbruck, Austria.
authors.forename: F F
authors.initials: FF
authors.lastname: Moebius
authors.forename: G G
authors.initials: GG
authors.lastname: Burrows
authors.forename: J
authors.initials: J
authors.lastname: Striessnig
authors.forename: H
authors.initials: H
authors.lastname: Glossmann
mesh_terms: D000345:Affinity Labels; D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D005260:Female; D006168:Guinea Pigs; D066298:In Vitro Techniques; D007511:Ischemia / Q000188:drug therapy*; D008099:Liver / Q000378:metabolism*; D008297:Male; D008565:Membrane Proteins / Q000302:isolation & purification / Q000378:metabolism*; D008970:Molecular Weight; D010777:Photochemistry; D011869:Radioligand Assay; D017480:Receptors, sigma / Q000378:metabolism; D012995:Solubility; D014700:Verapamil / Q000031:analogs & derivatives / Q000378:metabolism
chemical_list: D000345:Affinity Labels; D008565:Membrane Proteins; D017480:Receptors, sigma; D014700:Verapamil; C052752:emopamil