PRIMARY STRUCTURE OF PROSTAGLANDIN-G/H SYNTHASE FROM SHEEP VESICULAR GLAND DETERMINED FROM THE COMPLEMENTARY-DNA SEQUENCE
PRIMARY STRUCTURE OF PROSTAGLANDIN-G/H SYNTHASE FROM SHEEP VESICULAR GLAND DETERMINED FROM THE COMPLEMENTARY-DNA SEQUENCE
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DOI：
10.1073/pnas.85.5.1412
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发表时间：
1988-03-01
影响因子：
11.1
通讯作者：
SMITH, WL
中科院分区：
文献类型：
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作者：
DEWITT, DL;SMITH, WL
前列腺素G/H合酶（8，11，14-二十碳三烯酸酯，氢供体：氧氧化还原酶，EC 1.14.99.1）催化花生四烯酸转化为前列腺素内过氧化物G和H的形成中的第一步，花生四烯酸转化为前列腺素内过氧化物G和H。这种酶是非甾体抗炎药的作用部位。我们已经分离出一个2.7-脱氢酶互补DNA（cDNA），包括整个编码区的前列腺素G/H合成酶从绵羊水泡腺。该cDNA克隆自由泡状腺的poly（A）+ RNA制备的cDNA gt 10文库，与单一的2.75-腺苷酸酶mRNA种类杂交。使用从纯化的酶制备的胰蛋白酶肽的氨基酸序列建模的寡核苷酸探针选择cDNA克隆。全长cDNA编码600个氨基酸的蛋白质，包括24个氨基酸的信号序列。将cDNA鉴定为编码前列腺素G/H合酶是基于将包含103个氨基酸的7种肽的氨基酸序列与从cDNA的核苷酸序列推导的氨基酸序列进行比较。缺乏信号肽的未糖基化酶的分子量为65，621。该合成酶是一种糖蛋白，有三个潜在的N-糖基化位点，其中两个位于分子的氨基末端。据报道被阿司匹林乙酰化的丝氨酸位于羧基末端附近的530位。合成酶的序列与氨基酸或核苷酸序列文库中的任何其他蛋白质的序列之间没有显著的相似性，并且从氨基酸序列中不明显存在血红素结合位点。编码前列腺素G/H合酶的全长cDNA克隆的可用性应有助于研究这种酶的表达调节以及对催化和与抗炎药物相互作用重要的结构特征。
Prostaglandin G/H synthase (8,11,14-icosatrienoate, hydrogen-donor:oxygen oxidoreductase, EC 1.14.99.1) catalyzes the first step in the formation of prostaglandins and thromboxanes, the conversion of arachidonic acid to prostaglandin endoperoxides G and H. This enzyme is the site of action of nonsteroidal anti-inflammatory drugs. We have isolated a 2.7-kilobase complementary DNA (cDNA) encompassing the entire coding region of prostaglandin G/H synthase from sheep vesicular glands. This cDNA, cloned from a .lambda.gt10 library prepared from poly(A)+ RNA of vesicular glands, hybridizes with a single 2.75-kilobase mRNA species. The cDNA clone was selected using oligonucleotide probes modeled from amino acid sequences of tryptic peptides prepared from the purified enzyme. The full-length cDNA encodes a protein of 600 amino acids, including a signal sequence of 24 amino acids. Identification of the cDNA as coding for prostaglandin G/H synthase is based on comparison of amino acid sequences of seven peptides comprising 103 amino acids with the amino acid sequence deduced from the nucleotide sequence of the cDNA. The molecular weight of the unglycosylated enzyme lacking the signal peptide is 65,621. The synthase is a glycoprotein, and there are three potential sites for N-glycosylation, two of them in the amino-terminal half of the molecule. The serine reported to be acetylated by aspirin is at position 530, near the carboxyl terminus. There is no significant similarity between the sequence of the synthase and that of any other protein in amino acid or nucleotide sequence libraries, and a heme binding site(s) is not apparent from the amino acid sequence. The availability of a full-length cDNA clone coding for prostaglandin G/H synthase should facilitate studies of the regulation of expression of this enzyme and the structural features important for catalysis and for interaction with anti-inflammatory drugs.