Catalysis by Prostaglandin Endoperoxide H Synthases

描述（由申请人提供）：前列腺素内过氧化物H合成酶-1和-2 (PGHS-1和-2)，也称为环氧合酶-1和-2 (COX-1和COX-2)，在前列腺素（PG）生物合成的承诺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，COX抑制剂包括非特异性非甾体抗炎药（nsnsaaids）和COX-2特异性抑制剂coxib。COX抑制剂是美国使用最广泛的药物，然而，使用这些抑制剂具有显著的风险。每年约有20 000人死于这些药物的不良影响，其分子基础尚不清楚。使用纯化的人类PGHS，我们发现PGHS活性通过一种不寻常的变构机制被所有常见的脂肪酸（FAs）调节，包括那些不是PGHS底物的脂肪酸。FAs可以刺激或抑制PGHS活性，具体作用取决于PGHS异构体和FA。这种效应发生在生理FA浓度下，并在细胞中以及纯化酶中观察到。FAs调控PGHS的生化基础涉及组成PGHS同型二聚体的单体之间的串扰。虽然单体具有相同的氨基酸序列，但组成PGHS同型二聚体的两个单体的构象不同。一个单体结合FAs并作为变构单体，而另一个作为催化单体。最后也是重要的是，纯化的人PGHS对COX抑制剂的反应是由FAs调节的，同样取决于FA、抑制剂和PGHS异构体。对于不同的COX抑制剂，FAs可以影响抑制剂与一种或两种单体的结合。本研究的目的是确定pghs及其对广泛使用的COX抑制剂的反应如何在分子、细胞和全动物水平上受到FAs的影响。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酶所在的FA环境（FA tone）的显著调节，并且这种FA环境受到饮食中FA组成的重要影响。我们假设每个人在遗传背景下的饮食习惯都会导致FA音调。我们推测，某些FA音调使易感个体易患COX抑制剂的不良后果。我们期望我们的研究描述FA/COX抑制剂的相互作用将是改变COX抑制剂对不同饮食人群的处方方式的第一步，并调整饮食以提供COX抑制剂更安全的使用。

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. PUBLIC HEALTH RELEVANCE: This research is relevant to public health because it will provide a fundamental understanding of how different kinds of dietary fats can alter responses to common anti- inflammatory and analgesic drugs including aspirin, celecoxib, ibuprofen and naproxen. These drugs cause many adverse cardiovascular, renal and gastrointestinal side effects. Understanding how differences in the fat content of the diet can alter responses to these drugs will lead to changes in the way they are prescribed to people on different diets and will minimize the side effects.