Limon I, Coupry I, Lanier S M, Parini A
Centre National de la Recherche Scientifique URA 1482, Faculté de Médecine Necker-Enfants Malades, Paris, France.
J Biol Chem. 1992 Oct 25;267(30):21645-9.
The imidazoline-guanidinium receptive site (IGRS) is a membrane-bound protein that may mediate some of the pharmacological effects of imidazoline and guanidinium compounds. The structure and functionality of this protein are unknown but, in addition to its location at the plasma membrane, it is found in high density in the outer membrane of mitochondria (Tesson, F., Prip-Buus, C., Lemoine, A., Pegorier, J.-P., and Parini, A. (1991) J. Biol. Chem. 266, 155-160). Using a two-step procedure, we report the purification of mitochondrial IGRS from rabbit kidney to the apparent homogeneity. After solubilization of mitochondrial membranes with digitonin, an apparently homogeneous IGRS preparation was obtained by two sequential purification steps, chromatofocusing and hydroxylapatite-agarose chromatography. One- and two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of the purified preparation after silver staining or radioiodination indicated that IGRS binding subunit was purified at the apparent homogeneity since a single band (M(r) approximately 60,000) was observed. IGRS behaves as an acidic protein (pI 5.5) whose binding activity is regulated by H+ concentration near a physiological pH of 7.4. The ability to achieve rapid purification of IGRS should facilitate efforts to define molecular properties and functionality of this protein.
咪唑啉 - 胍基受体位点（IGRS）是一种膜结合蛋白，可能介导咪唑啉和胍基化合物的一些药理作用。该蛋白的结构和功能尚不清楚，但除了位于质膜外，还在线粒体外膜中高浓度存在（Tesson, F., Prip-Buus, C., Lemoine, A., Pegorier, J.-P., and Parini, A. (1991) J. Biol. Chem. 266, 155 - 160）。我们采用两步法报道了从兔肾中纯化线粒体IGRS至表观均一性。用洋地黄皂苷溶解线粒体膜后，通过连续的两步纯化步骤，即色谱聚焦和羟基磷灰石 - 琼脂糖色谱法，获得了表观均一的IGRS制剂。对纯化制剂进行银染或放射性碘化后的一维和二维十二烷基硫酸钠 - 聚丙烯酰胺凝胶电泳分析表明，IGRS结合亚基已纯化至表观均一性，因为只观察到一条带（相对分子质量约为60,000）。IGRS表现为一种酸性蛋白（pI 5.5），其结合活性在生理pH 7.4附近受H⁺浓度调节。能够快速纯化IGRS应有助于确定该蛋白的分子特性和功能。
