Plant Storage Lipid Biosynthesis

植物贮藏脂质生物合成

• 批准号: RGPIN-2014-04585
• 负责人: Weselake, Randall
• 金额: $ 4.3万
• 依托单位: University of Alberta
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2017
• 资助国家: 加拿大
• 起止时间: 2017-01-01 至 2018-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=630285
• 关键词: Plant; Storage; Lipid; Biosynthesis

Rising incomes, population growth, and an urgent need for renewable liquid fuels are driving an unprecedented global demand for seed oils. This year it is estimated that Canada will produce about 16 million metric tonnes of canola generating about $19 billion in economic activity for the nation (Canola Council of Canada). The overall goal of my research program is to understand how seed oil formation is controlled as a basis for developing canola with increased seed oil content and oil that can be designated as “zero saturate” to increase both the competitiveness and marketability of the oil. In general, increased consumption of saturated fats tends to be associated with increased prevalence of cardiovascular disease. During seed development, a series of oil-building enzymes (biological catalysts) drive the formation of oil. The action of these enzymes can influence both the quantity of oil formed and the composition and properties of the oil. The proposed research program focuses on two types of enzymes that drive reactions in the oil-building pathway. One of these enzymes (abbreviated DGAT) drives the final step in oil formation whereas the other enzyme (abbreviated LACS) drives the formation of building blocks for use in the final oil-building step. Recombinant DNA technology will be used to modify canola DGAT and the resulting modified forms of the enzyme will undergo biochemical testing. Enzyme modification will involve removing sections from the enzyme or making specific changes within the enzyme. This type of probing should provide some insight into how the enzyme operates and is able to accommodate certain oil-building blocks. High throughput methods will be used to generate a multitude of enzyme variants, some of which may have the characteristics sought. Production and identification of functional DGAT variants will be conducted using a yeast system whereas a bacterial system will be used for this purpose in the case of LACS. DGAT variants which drive the formation of oils reduced in saturation will be selected for further biochemical testing. In the case of LACS, enzyme variants will be selected which drive the formation of oil-building blocks with reduced saturation. Promising DGAT or LACS variants will then be introduced into a model oilseed plant (thale cress), via genetic engineering, to see if the degree of saturation is reduced in the oil. Enzyme variants which reduce saturation in thale cress seed oil will then be evaluated for their ability to reduce saturation in canola oil. We will also attempt to purify, crystallize and determine the detailed structure of canola DGAT in order to find out what really “makes it tick”. If this high risk component of our work is successful, it will complement our work on DGAT modification. Currently, there is no detailed structural information available on this enzyme. Detailed information on the structure and action of this key oil-building enzyme will lead to the development of other strategies for modifying seed oil production. This plant oil formation research will rely on methods from numerous disciplines including plant biochemistry, structural biology, molecular breeding, molecular biology and genetic-engineering. The proposed research program will also contribute to the training of highly qualified personnel in the burgeoning area of plant oils research.

收入的增加、人口的增长以及对可再生液体燃料的迫切需求正在推动全球对种子油的需求达到前所未有的水平。据加拿大油菜籽委员会估计，今年加拿大将生产约 1600 万吨油菜籽，为国家带来约 190 亿美元的经济活动。我的研究计划的总体目标是了解如何控制种子油的形成，作为开发种子油含量增加的芥花籽油和可指定为“零饱和”的油的基础，以提高油的竞争力和适销性。一般来说，饱和脂肪摄入量的增加往往与心血管疾病患病率的增加有关。在种子发育过程中，一系列造油酶（生物催化剂）驱动油的形成。这些酶的作用可以影响形成的油的量以及油的成分和特性。拟议的研究计划重点关注两种在造油途径中驱动反应的酶。其中一种酶（缩写为 DGAT）驱动油形成的最后一步，而另一种酶（缩写为 LACS）则驱动用于最终油形成步骤的结构单元的形成。重组 DNA 技术将用于修饰油菜 DGAT，所得酶的修饰形式将接受生化测试。酶修饰将涉及从酶中去除部分或在酶内进行特定的改变。这种类型的探测应该可以深入了解酶的运作方式以及能够适应某些油构建块。高通量方法将用于产生多种酶变体，其中一些可能具有所寻求的特征。功能性 DGAT 变体的生产和鉴定将使用酵母系统进行，而在 LACS 的情况下，将使用细菌系统来实现此目的。将选择驱动饱和度降低的油形成的 DGAT 变体进行进一步的生化测试。就 LACS 而言，将选择酶变体来驱动饱和度降低的石油结构块的形成。然后，将通过基因工程将有前景的 DGAT 或 LACS 变体引入模型油籽植物（拟南芥）中，以观察油中的饱和度是否降低。然后将评估降低拟南芥种子油饱和度的酶变体降低菜籽油饱和度的能力。我们还将尝试纯化、结晶并确定双低油菜籽 DGAT 的详细结构，以找出真正“让它发挥作用”的因素。如果我们工作的这一高风险部分取得成功，它将补充我们在 DGAT 修改方面的工作。目前，还没有关于这种酶的详细结构信息。关于这种关键的造油酶的结构和作用的详细信息将有助于开发其他改变种子油生产的策略。这项植物油形成研究将依赖于植物生物化学、结构生物学、分子育种、分子生物学和基因工程等众多学科的方法。拟议的研究计划还将有助于培训新兴植物油研究领域的高素质人才。