Molecular biological and biochemical characterization of the human type 2 selenodeiodinase
Molecular biological and biochemical characterization of the human type 2 selenodeiodinase
复制标题
DOI：
10.1210/en.137.8.3308
复制
发表时间：
1996-08-01
期刊：
影响因子：
4.8
通讯作者：
Larsen, PR
中科院分区：
文献类型：
--
作者：
Salvatore, D;Bartha, T;Larsen, PR
2 型脱碘酶 (DZ) 是一种低 K-m 碘甲状腺原氨酸脱碘酶，可催化从 T-4 或 rT(3) 的酚环中去除单个碘。我们对 Genethon (Z44085) 从人类婴儿大脑 cDNA 文库中制备的部分互补 DNA (cDNA) 克隆（2.1 KB）的开放阅读框进行了测序和亚克隆。该开放阅读框编码推测的 31 kDa、由 273 个氨基酸组成的蛋白质，与 Rana catesbeiana D2 蛋白质的相似性超过 70%。 cDNA 的瞬时表达在 293-HEK 细胞中产生低 K-m（T-4 为 5 nM；rT(3) 为 8 nM）丙硫氧嘧啶和金硫葡萄糖抗性 5'-脱碘酶。人 D2 与人 1 型 (D1) 和 3 型 (D3) 脱碘酶一样，是一种硒酶，证据如下：1) 存在两个符合读框的 UGA 密码子（位置 133 和 266），2) 在 D2 cDNA 转染的细胞中合成 31-kDa Se-75 标记的蛋白质，以及 3) 需要3'-硒代半胱氨酸掺入序列元件用于其翻译。与 D1 和 D3 不同，我们无法用 N-溴乙酰基 [I-125]T-3 或 -T-4 共价标记过表达的 D2。我们发现人类 D2 信使 RNA 为 7-8 kilobase，在大脑、胎盘以及令人惊讶的心肌和骨骼肌中表达。 2 型脱碘酶活性也存在于人类骨骼肌中。这些结果表明 D2 具有区别于其他两种硒代脱碘酶的独特特征。 D2 在肌肉中的表达表明它可能在外周以及细胞内 T-3 的产生中发挥作用。
Type 2 deiodinase (DZ) is a low K-m iodothyronine deiodinase that catalyzes the removal of a single iodine from the phenolic ring of T-4 or rT(3). We sequenced and subcloned the open reading frame from a partial complementary DNA (cDNA) clone (2.1 kilobases) prepared by Genethon (Z44085) from a human infant brain cDNA library. The open reading frame encodes a putative 273-amino acid protein of 31 kDa with greater than 70% similarity to the Rana catesbeiana D2 protein. Transient expression of the cDNA produces a low K-m (5 nM for T-4; 8 nM for rT(3)) propylthiouracil- and gold thioglucose-resistant 5'-deiodinase in 293-HEK cells. Human D2, like human type 1 (D1) and type 3 (D3) deiodinases, is a selenoenzyme, as evidenced by 1) the presence of two in-frame UGA codons (positions 133 and 266), 2) the synthesis of a 31-kDa Se-75-labeled protein in D2 cDNA-transfected cells, and 3) the requirement for a 3'-selenocysteine incorporation sequence element for its translation. Unlike D1 and D3, we were not able to covalently label overexpressed D2 with N-bromoacetyl [I-125]T-3 or -T-4. We found that the human D2 messenger RNA is 7-8 kilobases and is expressed in brain, placenta, and, surprisingly, cardiac and skeletal muscle. Type 2 deiodinase activity was also present in human skeletal muscle. These results indicate that there are unique features of D2 that distinguish it from the two other seleno-deiodinases. The expression of D2 in muscle suggests that it could play a role in peripheral, as well as intracellular, T-3 production.