Funk C D, Funk L B, Kennedy M E, Pong A S, Fitzgerald G A
Division of Clinical Pharmacology, Vanderbilt University, Nashville, Tennessee 37232.
FASEB J. 1991 Jun;5(9):2304-12.
Platelets metabolize arachidonic acid to thromboxane A2, a potent platelet aggregator and vasoconstrictor compound. The first step of this transformation is catalyzed by prostaglandin (PG) G/H synthase, a target site for nonsteroidal antiinflammatory drugs. We have isolated the cDNA for both human platelet and human erythroleukemia cell PGG/H synthase using the polymerase chain reaction and conventional screening procedures. The cDNA encoding the full-length protein was expressed in COS-M6 cells. Microsomal fractions from transfected cells produced prostaglandin endoperoxide-derived products which were inhibited by indomethacin and aspirin. Mutagenesis of the serine residue at position 529, the putative aspirin acetylation site, to an asparagine reduced cyclooxygenase activity to barely detectable levels, an effect observed previously with the expressed sheep vesicular gland enzyme. Platelet-derived growth factor and phorbol ester differentially regulated the expression of PGG/H synthase mRNA levels in the megakaryocytic/platelet-like HEL cell line. The PGG/H synthase gene was assigned to chromosome 9 by analysis of a human--hamster somatic hybrid DNA panel. The availability of platelet PGG/H synthase cDNA should enhance our understanding of the important structure/function domains of this protein and its gene regulation.
血小板将花生四烯酸代谢为血栓素A2，这是一种强效的血小板聚集剂和血管收缩剂化合物。这种转化的第一步由前列腺素（PG）G/H合酶催化，该酶是非甾体抗炎药的作用靶点。我们使用聚合酶链反应和传统筛选程序分离出了人血小板和人红白血病细胞PGG/H合酶的cDNA。编码全长蛋白的cDNA在COS-M6细胞中表达。转染细胞的微粒体部分产生前列腺素内过氧化物衍生产物，这些产物受到吲哚美辛和阿司匹林的抑制。将第529位丝氨酸残基（推测的阿司匹林乙酰化位点）突变为天冬酰胺，可使环氧化酶活性降低到几乎检测不到的水平，这一效应先前在表达的绵羊精囊腺酶中也观察到。血小板衍生生长因子和佛波酯对巨核细胞样/血小板样HEL细胞系中PGG/H合酶mRNA水平的表达有不同的调节作用。通过对人-仓鼠体细胞杂交DNA面板的分析，将PGG/H合酶基因定位到9号染色体上。血小板PGG/H合酶cDNA的获得应能增强我们对该蛋白重要结构/功能域及其基因调控的理解。
