GLYCEROLIPID METABOLISM IN MICROBODIES (PEROXISOMES)

微生物（过氧化物酶体）中的甘油脂代谢

• 批准号: 2379573
• 负责人: AMIYA K HAJRA
• 金额: $ 23.47万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 1979
• 资助国家: 美国
• 起止时间: 1979-12-01 至 1999-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2379573
• 关键词: acyl coA; acyltransferase; aldehyde /ketone oxidoreductase; complementary DNA; dihydroxyacetone; enzyme biosynthesis; enzyme mechanism; enzyme structure; glycerol; guinea pigs; human tissue; immunologic assay /test; inborn lipid /lipoprotein disorder; inborn metabolism disorder; laboratory mouse; laboratory rat; lipid biosynthesis; membrane biogenesis; membrane lipids; molecular cloning; oxidoreductase; peroxisome; protein purification; protein structure function; thioester; tissue /cell culture

Microbodies, i.e., peroxisomes and microperoxisomes, are ubiquitous catalase-containing subcellular organelles whose exact function in cellular metabolism is not clear. It is generally believed that the major function of these organelles is to metabolize various compounds by oxidation. However, we have shown that in animals these organelles also play a major role in membrane lipid biosynthesis. We demonstrated that the key enzymes of the acyl dihydroxyacetone phosphate pathway for lipid biosynthesis are localized in the membrane of microbodies. Thus pathway is the only route for the biosynthesis of either lipids (glycerol ether lipids and plasmalogens), which are present in all animals. The obligatory role of peroxisomes in ether lipid biosynthesis is manifested by the discovery of a number of genetic diseases (Zellweger cerebrohepatorenal syndrome, neonatal adrenoleukodystrophy, rhizomelic chondrodysplasia punctata, etc.) in which tissues of the patients are deficient in both peroxisomes and ether lipids. We have shown that the absence of ether lipids is due to the deficiency of a key peroxisomal enzyme which catalyzes the synthesis of acyl dihydroxyacetone phosphate. The major objective of this research proposal is to investigate the role of microbodies in the regulation of cellular lipid biogenesis at the molecular level. We will pursue our hypothesis that the main function of microbodies is to compartmentalize the substrates with specific topologic arrangements of the enzymes so that biochemical reactions leading to the formation of membrane lipid intermediates can be effectively regulated. The studies on ate purification and properties of peroxisomal membrane-bound acyl DHAP pathway enzymes and the determination of their molecular structures will be continued. The following specific aims are proposed: 1) To purify the two key enzymes of the acyl DHAP pathway, dihydroxyacetone phosphate (DHAP) acyltransferase and acyl/alkly DHAP reductase, with modified protocols so that sufficient quantities are obtained from structural studies and antibodies production. The molecular properties, regulation and biogenesis of these two enzymes will be investigated by determining their primary structure by cloning and sequencing the corresponding c-DNA's. 2) To generate specific antibodies against these two enzymes and to use these antibodies to study the tissue distribution, subcellular localization and regulation of the enzymes at the molecular level. These antibodies will be also used to detect the presence of mutant enzyme in peroxisomal genetic diseases such as Zellweger syndrome and rhizomelic chondrodysplasta punctata. 3) To study the mechanism of enzymatic generation and transport of metabolites for lipid biosynthesis in microbodies. The biochemical mechanism for the synthesis of specific membrane lipids, such as ethanolamine plasmalogens, from the intermediates of the acyl DHAP pathway will be investigated. 4). To investigate the molecular structure, function and regulation of a peroxisomal lipid-metabolizing enzyme, acyl Coenzyme A thioesterase, a very active constitutive brain enzyme which is also highly induced in rodent liver by peroxisome-proliferating drugs.

微型生物，即过氧化物酶体和微氧酶体，无处不在 含过氧化氢酶的亚细胞细胞器，其精确功能在细胞中 代谢尚不清楚。 通常认为主要功能 这些细胞器中是通过氧化代谢各种化合物。 但是，我们已经表明，在动物中，这些细胞器也发挥了作用 在膜脂质生物合成中的作用。 我们证明了关键酶 用于脂质生物合成的酰基二羟基丙酮磷酸途径是 位于微生物膜中。 因此，途径是唯一的路线 对于两种脂质的生物合成（甘油乙醚脂质和 浆元），所有动物中都存在。 强制性的作用 醚脂质生物合成中的过氧化物酶体的发现表现出来 许多遗传疾病（Zellweger小脑脑炎综合征， 新生儿肾上腺素肌营养不良，根茎软骨发育症状等） 其中患者的组织在过氧化物酶体和 醚脂质。 我们已经表明，没有醚脂质是由于 关键过氧化物酶体酶的缺乏，该酶催化合成 酰基二羟基丙酮磷酸盐。 该研究建议的主要目的是调查 细胞脂质生物发生调节的微生物在分子上 等级。 我们将提出假设，即微生物的主要功能 是通过特定的拓扑布置将基材分开 酶的生化反应导致形成 膜脂质中间体可以有效调节。 研究 过氧化物酶体结合的酰基DHAP的纯化和特性 途径酶及其分子结构的测定将是 持续。 提出了以下特定目标：1）净化两者 酰基DHAP途径的关键酶，二羟基丙酮磷酸盐（DHAP） 酰基转移酶和酰基/碱DHAP还原酶，具有改良的方案 从结构研究获得了足够的数量， 抗体生产。 分子特性，调节和生物发生 在这两种酶中，将通过确定其主要 通过克隆和测序相应的C-DNA来结构。 2）到 生成针对这两种酶的特定抗体并使用这些抗体 研究组织分布，亚细胞定位和 调节分子水平的酶。 这些抗体将是 还用于检测过氧化物酶体遗传中突变酶的存在 Zellweger综合征和根茎软骨塑料等疾病 点状。 3）研究酶产生和运输的机制 微生物中脂质生物合成的代谢产物。 生化 合成特定膜脂质的机制，例如 乙醇胺浆元，来自酰基DHAP途径的中间体 将被调查。 4）。 为了研究分子结构， 过氧化物酶体脂质代谢酶的功能和调节酰基，酰基 辅酶A硫酯酶，一种非常活跃的本构脑酶，是 也通过过氧化物酶体增殖药物高度诱导啮齿动物肝脏。