DeWitt D L, Smith W L
Department of Biochemistry, Michigan State University, East Lansing 48824.
Proc Natl Acad Sci U S A. 1988 Mar;85(5):1412-6. doi: 10.1073/pnas.85.5.1412.
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 gt 10 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.
前列腺素G/H合酶（8,11,14-二十碳三烯酸，氢供体：氧氧化还原酶，EC 1.14.99.1）催化前列腺素和血栓素形成的第一步，即花生四烯酸转化为前列腺素内过氧化物G和H。该酶是非甾体抗炎药的作用位点。我们从绵羊精囊腺中分离出了一个2.7千碱基的互补DNA（cDNA），它包含前列腺素G/H合酶的整个编码区。这个cDNA是从用精囊腺的聚腺苷酸加尾RNA构建的λgt 10文库中克隆得到的，它与一种单一的2.75千碱基的mRNA杂交。该cDNA克隆是使用根据从纯化酶制备的胰蛋白酶肽段的氨基酸序列构建的寡核苷酸探针筛选出来的。全长cDNA编码一个600个氨基酸的蛋白质，包括一个24个氨基酸的信号序列。将该cDNA鉴定为编码前列腺素G/H合酶是基于将由7个肽段组成的103个氨基酸的氨基酸序列与从cDNA的核苷酸序列推导出来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。该合酶是一种糖蛋白，有三个潜在的N-糖基化位点，其中两个在分子的氨基末端一半区域。据报道被阿司匹林乙酰化的丝氨酸位于第530位，靠近羧基末端。在氨基酸或核苷酸序列文库中，该合酶的序列与任何其他蛋白质的序列在氨基酸或核苷酸序列上均无明显相似性，并且从氨基酸序列中也看不出有血红素结合位点。编码前列腺素G/H合酶的全长cDNA克隆的获得应有助于对该酶表达调控以及对催化作用和与抗炎药相互作用重要的结构特征的研究。
