Understanding the roles of diacylglycerol acyltransferases in triacylglycerol biosynthesis and storage

了解二酰甘油酰基转移酶在三酰甘油生物合成和储存中的作用

• 批准号: RGPIN-2017-04355
• 负责人: Stone, Scot
• 金额: $ 4.95万
• 依托单位: University of Saskatchewan
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2022
• 资助国家: 加拿大
• 起止时间: 2022-01-01 至 2023-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=744103
• 关键词: Understanding; roles; diacylglycerol; acyltransferases; triacylglycerol

Triacylglycerol (TG) is quantitatively the most important storage form of metabolic energy for most living organisms. TGs consist of a glycerol backbone to which three fatty acids are attached. Because they are highly reduced and anhydrous, TGs are capable of providing much more energy than carbohydrate or protein. Eukaryotic organisms store TGs in cytosolic lipid droplets that can be metabolized during periods of nutrient deprivation when energy requirements are not being satisfied. Plants store large quantities of TGs in seeds that are used as energy for seedling growth and development. TG is synthesized by an enzyme called diacylglycerol acyltransferase (DGAT) which uses diacylglycerol and fatty acyl CoA as substrates. Two DGAT enzymes that share no sequence similarity have now been identified: DGAT1 and DGAT2. Although these two enzymes catalyze the same biochemical reaction in vitro, they appear to not have redundant functions in cells. DGAT1 and DGAT2 are proposed to produce distinct pools of TG. Both enzymes are present in the endoplasmic reticulum in cells; however, DGAT2 is unique in that it also translocates to lipid droplets, the organelle in which TG is stored. The presence of DGAT2 on lipid droplets is thought to allow for localized TG synthesis, leading to lipid droplet growth.Little is known about how the DGAT enzymes function in cells to promote the synthesis and storage of TG. Therefore, an integrated approach using biochemical and molecular techniques in parallel with proteomics and lipidomics will be used to understand how DGAT1 and DGAT2 work to synthesize TG. The specific objectives of the proposed research program are to: (1) Identify the lipid droplet targeting/binding domain of DGAT2, (2) Characterize the fatty acid signals involved in protein translocation to lipid droplets, (3) Determine if DGAT1 and DGAT2 produce distinct pools of TGs, and (4) Compare and contrast the enzymatic properties of DGAT1 and DGAT2 using a yeast expression system. The proposed studies will address fundamental questions in lipid metabolism and provide new insights into the molecular mechanism responsible for TG synthesis that may be of interest to both the pharmaceutical and agricultural industries. Due to the alarming increase in the prevalence of obesity in Canada, there is intense interest of pharmaceutical companies in Canada to develop therapeutic compounds to reduce TG synthesis and storage. In plants, TG is found in large quantities in oil seeds such as canola which represents a valuable resource for dietary consumption and industrial uses in Canada. Since many of the pathways for TG synthesis are conserved from mammals to plants, there is also the potential for knowledge transfer of our research to the agricultural sector with the goal of modifying TG production in oil seed crops. This could have a significant impact on plant biotechnology and the agricultural industry in Canada.

三酰甘油（TG）是大多数生物体代谢能量的最重要的储存形式。TG由三个脂肪酸连接的甘油主链组成。因为它们是高度还原和无水的，TG能够提供比碳水化合物或蛋白质更多的能量。真核生物将TG储存在胞质脂滴中，当能量需求得不到满足时，这些脂滴可以在营养缺乏期间代谢。植物在种子中储存大量TG，用作幼苗生长和发育的能量。 TG由称为二酰基甘油酰基转移酶（DGAT）的酶合成，其使用二酰基甘油和脂肪酰基CoA作为底物。现在已经鉴定出两种没有序列相似性的DGAT酶：DGAT 1和DGAT 2。虽然这两种酶在体外催化相同的生化反应，但它们在细胞中似乎没有多余的功能。DGAT 1和DGAT 2被认为产生不同的TG库。这两种酶都存在于细胞的内质网中;然而，DGAT 2的独特之处在于它也易位到脂滴，脂滴是储存TG的细胞器。脂滴上DGAT 2的存在被认为允许局部TG合成，导致脂滴生长。关于DGAT酶在细胞中如何发挥作用以促进TG的合成和储存，人们知之甚少。因此，将使用生物化学和分子技术与蛋白质组学和脂质组学并行的综合方法来了解DGAT 1和DGAT 2如何合成TG。 该研究计划的具体目标是：（1）鉴定DGAT 2的脂滴靶向/结合结构域，（2）表征蛋白质转运至脂滴中所涉及的脂肪酸信号，（3）确定DGAT 1和DGAT 2是否产生不同的TG库，以及（4）使用酵母表达系统比较和对比DGAT 1和DGAT 2的酶性质。拟议的研究将解决脂质代谢中的基本问题，并提供新的见解，负责TG合成的分子机制，可能是感兴趣的制药和农业行业。由于加拿大肥胖症患病率的惊人增加，加拿大制药公司对开发治疗化合物以减少TG合成和储存有浓厚的兴趣。在植物中，TG大量存在于油菜籽中，油菜籽是加拿大饮食消费和工业用途的宝贵资源。由于TG合成的许多途径从哺乳动物到植物都是保守的，因此我们的研究也有可能将知识转移到农业部门，目的是改变油料作物中的TG生产。这可能对加拿大的植物生物技术和农业产生重大影响。