Molecular Analysis of Plant Triacylglycerol Biosynthetic Enzymes

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Canola oil is made in the seed of the canola plant Brassica napus. Besides being a valuable food source, canola oil is part of a large agricultural industry that contributes to Canada's economy over 19 billion dollars annually. Our long term goals are to enhance the oil quantity and quality in the canola plant. Proteins located in the membranes of cells make canola oil. These proteins are enzymes that work together to make fats including triacyglycerol (TAG), a predominant fat in canola oil. In this research program, the last protein in pathway toward TAG production, Brassica napus DGAT1, will be studied in detail to gain insight into its function. To understand precisely how TAG, the main fat in canola, is generated, we require a molecular understanding of the protein that makes canola oil. Our research lab has expertise in determining the three-dimensional structure of proteins. A structure will allow us to examine the part(s) of the protein that carries out the function of producing TAG. In order to generate large quantities, we have expressed the canola plant DGAT1 protein in yeast. With this protein, we have determined that it produces fats similar to that found in the canola plant. This protein will be crystallized in order to see its three-dimensional structure. Its structural determination will require the use of X-ray exposure to the crystals. Structures will be sought both in the absence and presence of fat substrates. Once we can see the overall shape and details of the protein involved in canola oil production, we can then design protein that makes more canola oil or has preferences for unsaturated fat, a desired quality in food industry. This more efficient DGAT1 protein will be engineered in the canola plant, which will be assessed for canola oil production. Determining the structure of other TAG producing enzymes will also be conducted with the goals of influencing TAG production in canola plants. Future studies will focus on other enzymes in the TAG biosynthetic pathway. Overall, this study has the potential to improve the canola oil production in the seed of the canola plant, resulting in enhanced yield in the canola crops. This would have a strong effect on the canola crop industry and a strong economic advantage for Canada. This project will also contribute to the training of highly qualified personnel.