title: Purification and partial sequencing of high-affinity progesterone-binding site(s) from porcine liver membranes.
abstract: High-affinity progesterone-binding sites have been identified, characterized in and purified from porcine liver membranes. They were functionally solubilized by the non-denaturing zwitterionic detergent 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonic acid (Chaps, 20 mM. detergent/protein mass ratio 4:1) at a yield of 75-80%. Using [3H]progesterone as radioligand, binding studies showed high-affinity and low-affinity binding sites in microsomal preparations with an apparent Kd2 of 11 nM and an apparent Kd2 of 286 nM. In solubilized fractions the high-affinity binding sites were present at an apparent Kd2 of 69 nM. In both preparations, progesterone binding was time-dependent, saturable, reversible, and showed a similar hierachy of affinities for related steroids. A purification scheme was developed based on anion-exchanger procedures. The purified fraction as identified by maximum specific progester-one-binding activity contained two major polypeptides of apparent molecular masses (SDS/PAGE) of 28 kDa and 56 kDa, respectively. Sequencing of both polypeptides showed an identical amino terminus without significant identity in the amino acid sequence to any known protein primary structure.
authors.affiliation: Division of Clinical Pharmacology, Klinikum Innenstadt, University of Munich, Germany.
authors.forename: C
authors.initials: C
authors.lastname: Meyer
authors.forename: R
authors.initials: R
authors.lastname: Schmid
authors.forename: P C
authors.initials: PC
authors.lastname: Scriba
authors.forename: M
authors.initials: M
authors.lastname: Wehling
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D008862:Microsomes, Liver / Q000737:chemistry* / Q000378:metabolism; D008969:Molecular Sequence Data; D011374:Progesterone / Q000378:metabolism*; D011980:Receptors, Progesterone / Q000302:isolation & purification* / Q000378:metabolism; D017421:Sequence Analysis; D012995:Solubility; D013552:Swine
chemical_list: D011980:Receptors, Progesterone; D011374:Progesterone