Náray-Fejes-Tóth A, Fejes-Tóth G
Department of Physiology, Dartmouth Medical School, Lebanon, New Hampshire 03756, USA.
Endocrinology. 1995 Jun;136(6):2579-86. doi: 10.1210/endo.136.6.7750480.
11 beta-hydroxysteroid dehydrogenase (11 beta-OHSD) is thought to confer aldosterone specificity to mineralocorticoid target cells by protecting the mineralocorticoid receptor from occupancy by endogenous glucocorticoids. We have recently described a novel isoform of 11-OHSD in the renal aldosterone target cells (11 beta-OHSD/CD) that differs from the previously characterized isoform (11 beta-OHSD-1). Unlike 11-OHSD-1, the collecting duct enzyme catalyzes irreversible dehydrogenation, has a very high affinity for its substrate, and is tissue-specific. We report here the isolation, sequence, and characterization of a complementary DNA (cDNA) encoding the rabbit collecting duct 11 beta-OHSD/CD or 11 beta-OHSD type 2. The cDNA, isolated using expression screening in Xenopus oocytes, is 1.9 kilobases in length and encodes a protein of 406 amino acids with a predicted molecular mass of 44,130 daltons. The cloned enzyme has a Michaelis constant (Km) for corticosterone of 6.6 +/- 3 nM, catalyzes exclusively dehydrogenation, and uses only NAD as cofactor. The cloned enzyme shows 85% and 75% amino acid identity to the recently cloned human type 2 11 beta-OHSD and sheep kidney 11 beta-OHSD, respectively, whereas the overall homology to rat liver 11 beta-OHSD-1 is less than 20% The messenger RNA for this 11 beta-OHSD is expressed at very high levels in the renal collecting duct and at much lower levels in the colon. The intrarenal distribution was determined by reverse-transcription polymerase chain reaction in isolated nephron segments or cell types. The messenger RNA is present only in aldosterone target cells within the kidney, at highest levels in principal cells, at lower levels in intercalated cells, and in inner medullary cells. These data suggest that the 11 beta-OHSD cDNA from rabbit collecting duct cells encodes the enzyme that confers aldosterone selectivity to mineralocorticoid target cells.
11β-羟基类固醇脱氢酶（11β-OHSD）被认为通过保护盐皮质激素受体不被内源性糖皮质激素占据，从而赋予盐皮质激素靶细胞醛固酮特异性。我们最近在肾醛固酮靶细胞中描述了一种新型的11-OHSD同工型（11β-OHSD/CD），它与先前鉴定的同工型（11β-OHSD-1）不同。与11-OHSD-1不同，集合管酶催化不可逆的脱氢反应，对其底物具有非常高的亲和力，并且具有组织特异性。我们在此报告编码兔集合管11β-OHSD/CD或11β-OHSD 2型的互补DNA（cDNA）的分离、序列和特征。使用非洲爪蟾卵母细胞中的表达筛选分离出的该cDNA长度为1.9千碱基，编码一个由406个氨基酸组成的蛋白质，预测分子量为44,130道尔顿。克隆的酶对皮质酮的米氏常数（Km）为6.6±3 nM，仅催化脱氢反应，并且仅使用NAD作为辅因子。克隆的酶与最近克隆的人2型11β-OHSD和绵羊肾11β-OHSD分别具有85%和75%的氨基酸同一性，而与大鼠肝11β-OHSD-1的总体同源性小于20%。这种11β-OHSD的信使RNA在肾集合管中高水平表达，在结肠中表达水平低得多。通过逆转录聚合酶链反应在分离的肾单位节段或细胞类型中确定肾内分布。信使RNA仅存在于肾内的醛固酮靶细胞中，在主细胞中水平最高，在闰细胞和髓质内细胞中水平较低。这些数据表明，来自兔集合管细胞的11β-OHSD cDNA编码赋予盐皮质激素靶细胞醛固酮选择性的酶。
