Otterness D M, Weinshilboum R
Department of Pharmacology, Mayo Medical School, Rochester, MN 55905.
Chem Biol Interact. 1994 Jun;92(1-3):145-59. doi: 10.1016/0009-2797(94)90060-4.
Human tissues contain at least three well-characterized cytoplasmic sulfotransferase (ST) enzymes, dehydroepiandrosterone (DHEA) ST and two of phenol ST (PST). DHEA ST catalyzes the sulfation of DHEA and other steroids. We cloned and expressed two cDNAs for human liver DHEA ST. The cloning strategy involved the design of PCR primers directed against two conserved domains in ST proteins. These primers were used to generate a specific PCR product that was then used successfully to clone cDNAs for DHEA ST from a human liver cDNA library. Two cDNAs were isolated that were approximately 1.1 and 1.8 kb in length. These two clones had identical open reading frames. Both cDNAs produced enzymatically active DHEA ST protein in a mammalian expression system. Northern blot analysis confirmed the presence of 1.1 and 1.8 kb transcripts in human liver. cDNAs for a number of eukaryotic enzymes have now been cloned, and they share significant sequence homology. These ST cDNAs appear to fall into distinct groups on the basis of amino acid sequences of the proteins that they encode, thus demonstrating that the enzymes comprise a gene superfamily. We have also isolated, a genomic clone for human DHEA ST that contains approximately 3 kb of 5'-flanking sequence, exon 1 and 1.7 kb of intron 1. Characterization of the structure and regulatory elements of this gene should help to elucidate mechanisms involved in the regulation of DHEA ST in humans.
人体组织至少含有三种特征明确的细胞质硫酸转移酶（ST），即脱氢表雄酮（DHEA）ST和两种酚类ST（PST）。DHEA ST催化DHEA和其他类固醇的硫酸化反应。我们克隆并表达了人肝脏DHEA ST的两个cDNA。克隆策略涉及针对ST蛋白中两个保守结构域设计PCR引物。这些引物用于产生特定的PCR产物，然后成功地用于从人肝脏cDNA文库中克隆DHEA ST的cDNA。分离出两个长度分别约为1.1 kb和1.8 kb的cDNA。这两个克隆具有相同的开放阅读框。在哺乳动物表达系统中，这两个cDNA均产生具有酶活性的DHEA ST蛋白。Northern印迹分析证实人肝脏中存在1.1 kb和1.8 kb的转录本。现在已经克隆了许多真核酶的cDNA，它们具有显著的序列同源性。根据它们所编码蛋白质的氨基酸序列，这些ST cDNA似乎可分为不同的组，从而表明这些酶构成了一个基因超家族。我们还分离出了人DHEA ST的一个基因组克隆，它包含约3 kb的5'侧翼序列、外显子1和1.7 kb的内含子1。对该基因的结构和调控元件进行表征应有助于阐明人类DHEA ST调控所涉及的机制。
