Catalysis by Prostaglandin Endoperoxide H Synthases

描述(由申请人提供)：前列腺素内源性过氧化氢合成酶-1和-2(PGHS-1和-2)，也称为环氧合酶-1和环氧合酶-2(COX-1和COX-2)，在前列腺素(PG)生物合成的关键步骤中催化花生四烯酸(AA)转化为前列腺素H2(PGH2)。PGHS是COX抑制剂的主要靶点，包括非特异性非类固醇抗炎药(NsNSAIDs)和COX-2特异性抑制剂Coxibs。COX抑制剂是美国使用最广泛的药物。然而，这些抑制剂的使用有很大的风险。每年约有2万人死于这些药物的不良反应，其分子基础尚不清楚。使用纯化的人PGHS，我们发现PGHS的活性是通过一种不寻常的变构机制被所有常见的脂肪酸(FA)调节的，包括那些不是PGHS底物的脂肪酸。Fas可刺激或抑制PGHS活性，其具体作用依赖于PGHS异构体和FA。这种影响发生在生理浓度的FA中，并在细胞和纯化的酶中观察到。脂肪酸调节PGHS的生化基础是由PGHS同源二聚体组成的单体之间的串扰。虽然两个单体具有相同的氨基酸序列，但包含PGHS同源二聚体的两个单体的构象不同。一个单体与FAs结合，以变构单体的形式存在，而另一个单体则作为催化单体。最后，也是重要的是，纯化的人PGHS对COX抑制剂的反应受FAs的调节，同样依赖于FA、抑制剂和PGHS亚型。对于不同的COX抑制剂，FA可以影响抑制剂与一种或两种单体的结合。这项拟议研究的目标是在分子、细胞和整个动物水平上确定FAs如何影响PGHS及其对广泛使用的COX抑制剂的反应。我们的基本假设是，体内PG的产生和对COX抑制剂的反应都受到FA环境的显著调节，酶在其中发现自己的FA音调，并且这种FA环境受到饮食中FA成分的重要影响。我们推测，每个人在其遗传背景下，由于饮食习惯的影响，都会建立一个FA基调。我们推测，某些FA音调使易感个体容易受到COX抑制剂的不良后果的影响。我们预计，我们描绘FA/COX抑制剂相互作用的研究将是第一步，将导致改变给不同饮食的人开COX抑制剂的方式，并调整饮食，以提供更安全的COX抑制剂使用。

DESCRIPTION (provided by applicant): Prostaglandin endoperoxide H synthases-1 and -2 (PGHS-1 and -2), also known as cyclooxygenases-1 and -2 (COX-1 and COX-2), catalyze the conversion of arachidonic acid (AA) to prostaglandin H2 (PGH2) in the committed step of prostaglandin (PG) biosynthesis. PGHSs are the primary targets of COX inhibitors, which include nonspecific nonsteroidal anti-inflammatory drugs (nsNSAIDs) and COX-2 specific inhibitors called coxibs. COX inhibitors are the most widely used pharmaceutical agents in the U.S. However, the use of these inhibitors carries significant risks. About 20,000 deaths annually are attributable to adverse effects of these drugs, the molecular basis for which is unknown. Using purified human PGHSs, we have discovered that PGHS activities are modulated through an unusual allosteric mechanism by all common fatty acids (FAs) including those that are not PGHS substrates. FAs can stimulate or inhibit PGHS activity with the specific effect being dependent on the PGHS isoform and the FA. The effects occur at physiologic FA concentrations and are observed in cells as well as with purified enzymes. The biochemical basis for the regulation of PGHSs by FAs involves cross-talk between monomers comprising PGHS homodimers. Although the monomers have identical amino acid sequences, the conformations of the two monomers comprising a PGHS homodimer differ. One monomer binds FAs and behaves as an allosteric monomer while the other acts as the catalytic monomer. Finally and importantly, responses of purified human PGHSs to COX inhibitors are modulated by FAs, again depending on the FA, the inhibitor and the PGHS isoform. With different COX inhibitors, FAs can influence binding of an inhibitor to one or to both monomers. The goals of the proposed research are to determine how PGHSs and their responses to widely used COX inhibitors are affected by FAs at the molecular, cellular and whole animal levels. Our underlying hypothesis is that both in vivo PG production and responses to COX inhibitors are significantly modulated by the milieu of FAs in which the enzymes find themselves-the FA tone--and that this FA environment is importantly influenced by the FA composition of the diet. We presume that every individual establishes a FA tone as a consequence of dietary habits in the context of their genetic background. We speculate that certain FA tones predispose susceptible individuals to adverse consequences of COX inhibitors. We expect that our studies delineating FA/COX inhibitor interactions will be a first step leading to changes in the way COX inhibitors are prescribed to people on different diets and to dietary adjustments to provide for the safer use of COX inhibitors.