Roth G J, Siok C J, Ozols J
J Biol Chem. 1980 Feb 25;255(4):1301-4.
Prostaglandin synthetase contains both oxygenase and peroxidase activity and catalyzes the first step of prostaglandin synthesis. Aspirin (acetylsalicylic acid) inhibits oxygenase activity by acetylating a serine residue of the enzyme. In the current study, we have investigated the subunit structure of this complex enzyme and the stoichiometry of aspirin-mediated acetylation of the enzyme. The enzyme was purified to near homogeneity in both active and aspirin-acetylated forms. The purified protein was analyzed for enzymatic activity, [3H]acetate content following treatment with [acetyl-3H]aspirin, NH2-terminal sequence, and amino acid composition. The results show first, that the enzyme can be purified to near homogeneity in an active form; second, that the enzyme consists of a single polypeptide chain (molecular weight 72,000 by sodium dodecyl sulfate polyacrylamide gel electrophoresis) with a unique NH2-terminal sequence (Ala-Asp-Pro-Gly-Ala-Pro-Ala-Pro-Val-Asn-Pro-Met-Gly-); and third, that aspirin inhibits the enzyme by transfer of one acetate per enzyme monomer. Therefore, the two distinct enzymatic activities, oxygenation and peroxidation, are present in a single polypeptide chain. Experiments with a cross-linking agent indicate that in nonionic detergent the enzyme is a dimer of two identical subunits.
前列腺素合成酶兼具加氧酶和过氧化物酶活性，催化前列腺素合成的第一步。阿司匹林（乙酰水杨酸）通过使该酶的一个丝氨酸残基乙酰化来抑制加氧酶活性。在本研究中，我们研究了这种复合酶的亚基结构以及阿司匹林介导的该酶乙酰化的化学计量。该酶以活性形式和阿司匹林乙酰化形式均被纯化至接近均一状态。对纯化后的蛋白质进行了酶活性分析、用[乙酰-³H]阿司匹林处理后的[³H]乙酸含量分析、氨基末端序列分析和氨基酸组成分析。结果首先表明，该酶可以以活性形式纯化至接近均一状态；其次，该酶由一条单一多肽链组成（通过十二烷基硫酸钠聚丙烯酰胺凝胶电泳测得分子量为72,000），具有独特的氨基末端序列（丙氨酸-天冬氨酸-脯氨酸-甘氨酸-丙氨酸-脯氨酸-丙氨酸-脯氨酸-缬氨酸-天冬酰胺-脯氨酸-甲硫氨酸-甘氨酸-）；第三，阿司匹林通过每个酶单体转移一个乙酸基团来抑制该酶。因此，两种不同的酶活性，即加氧和过氧化活性，存在于一条单一多肽链中。用交联剂进行的实验表明，在非离子去污剂中该酶是由两个相同亚基组成的二聚体。
