Urade Y, Watanabe K, Eguchi N, Fujii Y, Hayaishi O
Department of Enzymes and Metabolism, Osaka Bioscience Institute, Japan.
J Biol Chem. 1990 Jul 15;265(20):12029-35.
9 alpha,11 beta-prostaglandin F2 was formed from prostaglandin D2 by its 11-ketoreductases in 100,000 x g supernatants of various bovine tissues in the presence of an NADPH-generating system. The reductase activities were high in liver (51.09 nmol/h/mg of protein), lung (24.99), and spleen (14.20); moderate in heart and pancreas (3.09-3.61); weak in stomach, intestine, colon, kidney, uterus, adrenal gland, and thymus (0.11-2.63); and undetectable in brain, retina, carotid artery, and blood (less than 0.10). No formation of prostaglandin F2 alpha from prostaglandin D2 was detected in all tissues. In immunotitration analyses with a polyclonal antibody specific for prostaglandin F synthetase, the reductase activities in lung and spleen showed identical titration curves to that of the purified synthetase and decreased to less than 15% of the initial activity under the condition of antibody excess. Prostaglandin F synthetase-immunoreactive protein in these two tissues showed peptide fingerprints identical to that of the purified enzyme after partial digestion with Staphylococcus aureus V8 protease. The antibody was partially cross-reactive to the reductase in liver (about 20% of that to the synthetase) but not to the reductase(s) in other tissues. The Km value for prostaglandin D2 of the reductase activity was the same in lung and spleen as that of the purified prostaglandin F synthetase (120 microM) but differed in liver (6 microM), heart, and pancreas (15 microM). The predominant distribution of prostaglandin F synthetase in lung and spleen was confirmed by radioimmunoassay (2.8 and 1.0 micrograms/mg protein, respectively) and Northern blot analyses. In immunoperoxidase staining, this enzyme was localized in alveolar interstitial cells and nonciliated epithelial cells in lung, histiocytes and/or dendritic cells in spleen, and a few interstitial cells in kidney and adrenal cortex.
在存在NADPH生成系统的情况下，9α,11β-前列腺素F2由前列腺素D2通过其11-酮还原酶在各种牛组织的100,000×g上清液中形成。还原酶活性在肝脏中较高（51.09 nmol/h/mg蛋白质），在肺中（24.99）和脾脏中（14.20）；在心脏和胰腺中中等（3.09 - 3.61）；在胃、肠、结肠、肾脏、子宫、肾上腺和胸腺中较弱（0.11 - 2.63）；在脑、视网膜、颈动脉和血液中未检测到（小于0.10）。在所有组织中均未检测到由前列腺素D2形成前列腺素F2α。在用针对前列腺素F合成酶的多克隆抗体进行的免疫滴定分析中，肺和脾脏中的还原酶活性显示出与纯化的合成酶相同的滴定曲线，并且在抗体过量的条件下降低至初始活性的不到15％。用金黄色葡萄球菌V8蛋白酶部分消化后，这两种组织中的前列腺素F合成酶免疫反应性蛋白显示出与纯化酶相同的肽指纹图谱。该抗体与肝脏中的还原酶有部分交叉反应（约为与合成酶交叉反应的20％），但与其他组织中的还原酶无交叉反应。肺和脾脏中还原酶活性对前列腺素D2的Km值与纯化的前列腺素F合成酶相同（120μM），但在肝脏中（6μM）、心脏和胰腺中（15μM）不同。通过放射免疫测定（分别为2.8和1.0μg/mg蛋白质）和Northern印迹分析证实了前列腺素F合成酶在肺和脾脏中的主要分布。在免疫过氧化物酶染色中，该酶定位于肺中的肺泡间质细胞和无纤毛上皮细胞、脾脏中的组织细胞和/或树突状细胞以及肾脏和肾上腺皮质中的一些间质细胞中。
