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MECHANISM OF FATTY ACID SYNTHESIS

脂肪酸合成机制

基本信息

批准号：
3225048
负责人：
JOHN J LUCAS
金额：
$ 15.34万
依托单位：
UPSTATE MEDICAL UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
1979
资助国家：
美国
起止时间：
1979-09-01 至 1993-12-31
项目状态：
已结题

项目摘要

The de novo synthesis of fatty acids from acetyl- and malonyl-CoA in animals is catalyzed by an enzyme comprised of two identical, multifunctional polypeptide subunits. The thioester substrates are loaded on a common 4'-phosphopantetheine site and the incorrectly loaded substrate is removed by reaction with coenzyme A through a kinetic self-editing mechanism. The covalently-attached acyl-intermediates are then transfered to sites for condensation, Beta-ketoacyl reduction, dehydration, enoyl reduction, and finally deacylation following completion of chain elongation. In the proposed study, a variety of techniques such as transient and steady-state kinetic analyses, equilibrium substrate binding, reactivity of thiol groups, hybridization and crosslinking will be employed to elucidate a detailed catalytic mechanism of this enzyme and its functional conformation states. Chemical modification with specific reagents will be carried out to detect and characterize amino-acid residues essential for activity. Experiments are designed to test for the apparent "half-of-the-sites" reactivity of 5,5 dithiobis (2,-nitrobenzoic acid), and for a requirement of specific subunit interactions for condensation and acyl-transfer between hydroxyl, pantetheine, and cysteine sites implicated by the recently postulated "head-to-tail" model. A "half-of-the-sites" model has been proposed by us to account for the unusual behavior of malic enzyme which supplies a portion of NADPH equivalents for fatty acid synthesis. The validity of this model will be examined by end group analysis, peptide mapping, and by determining the stoichiometry of catalytic sites and activity of matrix-bound monomeric units of the enzyme. Ultimately, results of this study will enhance our knowledge of lipid metabolism and provide a basis for the prevention, diagnosis, and treatment of disorders related to lipid metabolism such as artereosclerosis and obesity.

从乙酰辅酶A和丙二酰辅酶A从头合成脂肪酸动物是由一种酶催化的，多功能多肽亚基。将硫酯底物加载到在共同的4 ′-磷酸泛酰巯基乙胺位点上，通过动力学自编辑与辅酶A反应而被去除机制然后将共价连接的酰基中间体转移到到缩合、β-酮酰基还原、脱水、烯酰基还原，最后在链完成后脱酰伸长率在拟议的研究中，各种技术，如瞬态和稳态动力学分析，平衡底物结合，将采用硫醇基团的反应性、杂化和交联为了阐明该酶的详细催化机制，功能构象状态。化学改性与特定将进行试剂检测和表征氨基酸残留物活动必不可少。实验的目的是测试表面上的 5，5-二硫代双（2，-硝基苯甲酸）的“半位点”反应性，和对于缩合的特定亚基相互作用的要求，羟基、泛酰巯基乙胺和半胱氨酸位点之间的酰基转移最近提出的“头对尾”模型。我们提出了一个“一半的网站”模型来解释提供一部分NADPH的苹果酸酶的异常行为用于脂肪酸合成的当量。该模型的有效性将是通过末端基团分析、肽图谱分析和确定催化位点化学计量与基质结合单体的活性单位的酶。最终，这项研究的结果将提高我们的了解脂质代谢，为预防提供依据，诊断和治疗与脂质代谢有关的疾病，动脉硬化和肥胖。