Human heme oxygenase-2: characterization and expression of a full-length cDNA and evidence suggesting that the two HO-2 transcripts may differ by choice of polyadenylation signal.
Human heme oxygenase-2: characterization and expression of a full-length cDNA and evidence suggesting that the two HO-2 transcripts may differ by choice of polyadenylation signal.
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人血红素加氧酶-2：全长 cDNA 的表征和表达，以及表明两个 HO-2 转录物可能因聚腺苷酸化信号的选择而异的证据。
DOI：
10.1016/0003-9861(92)90481-b
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发表时间：
1992
影响因子：
3.9
通讯作者：
Maines,MD
中科院分区：
文献类型：
--
作者：
McCoubreyJr,WK;Ewing,JF;Maines,MD
我们通过 Northern blot 分析表明，人 HO-2 由两个转录本（1.3 和 1.7 kb）编码，并且通过 Southern blot 分析判断为单拷贝基因。我们进一步提供了基于 Northern 印迹和代表较大转录本的 cDNA 序列分析的证据，表明转录本在 3' 非翻译区有所不同。来自大鼠血红素加氧酶-2 基因的 274 个碱基对 DNA 片段（I. Cruse 和 M. D. Maines, 1988,J. Biol. Chem.263, 3348–3353）用于从 λgt11 中的胎儿肾文库中分离人 HO-2 cDNA。对命名为 hK-1 的克隆进行测序，确定 cDNA 插入片段长度为 1625 个碱基对，编码 313 个氨基酸的蛋白质。在 3' 非翻译区鉴定出两个由 440 个核苷酸分隔的共有聚腺苷酸化信号。 cDNA 插入片段的大小非常接近两个 mRNA 中较大的一个。核苷酸序列与预测编码区内的大鼠 HO-2 基因有 88% 的同一性，并且推定的翻译产物也估计与大鼠基因产物有 88% 的同一性 (M. O. Rotenberg 和 D. Maines, 1990, J. Biol. Chem.265, 7501)。预测的大小为 36 kDa，与通过蛋白质印迹分析在人睾丸微粒体中检测到的 HO2 非常吻合。此外，在大肠杆菌中表达的融合蛋白表现出显着的血红素加氧酶活性，该活性被Zn-和Sn-原卟啉（已知的真核血红素加氧酶抑制剂）抑制，但不被巯基试剂抑制。
We show by Northern blot analysis that human HO-2 is encoded by two transcripts (1.3 and 1.7 kb) and is a single-copy gene as judged by Southern blot analysis. We further provide evidence based on Northern blot and sequence analysis of a cDNA representing the larger transcript that the transcripts differ in the 3′ untranslated region. A 274-base-pair DNA fragment from the rat heme oxygenase-2 gene (I. Cruse and M. D. Maines, 1988,J. Biol. Chem.263, 3348–3353) was used to isolate a human HO-2 cDNA from a fetal kidney library in λgt11. The clone, designated hK-1, was sequenced and the cDNA insert was determined to be 1625 base pairs in length, encoding a protein of 313 amino acids. Two consensus polyadenylation signals separated by 440 nucleotides were identified in the 3′ untranslated region. The size of the cDNA insert closely approximated the larger of two mRNAs. The nucleotide sequence was 88% identical to the rat HO-2 gene within the predicted coding region and the putative translation product was also estimated to be 88% identical to the rat gene product (M. O. Rotenberg and D. Maines, 1990,J. Biol. Chem.265, 7501). The predicted size, 36 kDa, corresponded well with HO2 detected in human testis microsomes by Western blot analysis. Further, the fusion protein expressed inEscherichia colidisplayed significant heme oxygenase activity, which was inhibited by Zn- and Sn-protoporphyrins, known inhibitors of eukaryotic heme oxygenase, but not by sulfhydryl reagents.