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Researchers modify camelina to produce highest levels yet in transgenic plant oil of novel lipid acetyl-TAG; biofuel and industrial use

Researchers at Kansas State University led by Professor Timothy Durrett and their colleagues at Michigan State University and the University of Nebraska, Lincoln have engineered Camelina sativa—a non-food oilseed crop—to produce high levels (up to 85 mol%) of acetyl-triacylglycerols (acetyl-TAGs, or ac-TAGs)—a novel plant oil lipid with possible biofuel or industrial uses.

As reported in a paper in Plant Biotechnology Journal, this successful metabolic engineering and subsequent field production of the modified camelina crop marked the highest accumulation of the unusual oil achieved so far in transgenic plants. (Earlier work by Durrett and colleagues at the DOE Great Lakes Bioenergy Research Center had resulted in approximately a 60 mol% accumulation of ac-TAGs.)

Compared to most vegetable oils, acetyl-TAGs have reduced viscosity and improved cold temperature properties that confer advantages in applications as biodegradable lubricants, food emulsifiers, plasticizers, and ‘drop-in’ fuels for some diesel engines.

As one example, the viscosity of acetyl-TAG falls within or slightly above the specifications of diesel #4 fuel. By contrast, the higher viscosity of commercially produced vegetable oils prevents their direct use in most diesel engines; thus the vegetable oil needs either to be heated before entering the engine and fuel filters, or transesterified to methyl or ethyl esters to produce biodiesel.

As another example, acetyl-TAGs have markedly lower crystallization temperatures than the corresponding conventional TAG. The much lower crystallization temperature of acetyl-TAG will likely be reflected in improved pour point and cold filter plug points, the researchers said. Thus, these structures may provide improved low temperature performance for acetyl-TAG oils as lubricants, and fuels and also as alpha-tending emulsifiers in food applications.

Seed oils from nearly all plant species contain three acyl chains esterified to the three hydroxyl groups of glycerol. In major oilseed crops, five fatty acids (16:0, 18:0, 18:1, 18:2, and 18:3) predominate, but within the plant kingdom, over 200 ‘unusual’ oils occur in seeds, of which most are characterized by the presence of modified or uncommon fatty acids. These modifications include atypical chain length (C8–C22), unusual double bond positions, and addition of functional groups (e.g. hydroxyl, epoxy, etc.). Some of these modified structures have added-value properties that are currently used in a range of industrial applications.

3-acetyl-1,2-diacyl-sn-glycerols (acetyl-TAG) possess a particularly striking variation where the long-chain fatty acid at the sn-3 position of TAG is replaced by a two-carbon acetyl group.

—Liu et al. (2015a)

1-s2.0-S0926669014007067-gr1
Comparison of typical plant triacylglycerol structure with acetyl-triacylglycerol. The modified Camelina sativa plants produce 3-acetyl-1,2-diacyl-sn-glycerol in which an acetyl group replaces a long chain fatty acid at the sn-3 position. Liu et al. (2015a). Click to enlarge.

The presence of the sn-3 acetyl group confers useful physical, chemical and nutritional properties to these molecules. For example, they possess reduced kinematic viscosity and lower melting points compared to conventional triacylglycerols (here referred to aslcTAGs).

… Seeds of Euonymus alatus (Burning Bush) accumulate acetyl-TAGs as the major component of their storage oil. Previously, we identified an acetyltransferase, EaDAcT, from E. alatus that synthesizes acetyl-TAGs from DAG and acetyl-CoA in vitro and results in up to 45 mol% acetyl-TAG when expressed in wild-type Arabidopsis seeds.

… Here, we provide evidence that down-regulation of the DGAT1 pathway for synthesis of lcTAGs leads to the enhanced accumulation of acetyl-TAGs in Arabidopsis and Camelina. The successful application of this strategy to the oilseed crop Camelina sativa resulted in up to 85 mol% acetyl-TAGs in the oil, representing the highest levels of unusual lipids achieved in transgenic plants. The accumulation of these structurally different storage lipids by field-grown plants had minor or no impact on seed size, oil content, germination or oil mobilization by seedlings. In a concurrent study [Liu et al. 2015a] we obtained similar results from transformations of a high-oleic Camelina line. Analysis of acetyl-TAGs from field-grown plants demonstrated significant and useful differences in the physical properties of this unusual oil.

—Liu et al. (2015b)

The goal of Durrett’s research is to alter oilseeds to produce large amounts of modified oil that can be used as improved biofuels or in industrial and food-related applications. The research recently appeared in an open access paper in the journal Industrial Crops and Products and on the front cover of the Plant Biotechnology Journal.

Camelina can grow on poorer quality farmland, needs little irrigation or fertilizer, and produces seeds that can provide gallons of oil, Durrett said. It also can be rotated with wheat and could become a biofuel crop for semi-arid regions, including western Kansas and Colorado.

The camelina genome was recently sequenced, which has greatly helped Durrett and collaborators as they improve camelina’s oil properties to produce low-viscosity oil.

One of the team’s goals is to make commercial products using oils from the engineered plants. The researchers are analyzing these oils because their acetyl-TAGs possess unusual structures and have high value-added properties.

The basic problem is that most of our oilseed crops—such as canola or soybean—produce just a few fatty acids because we use them for nutritional needs. That’s great for a source of food, but makes doing any sort of chemistry more complicated.

—Timothy Durrett

The researchers think that camelina producing acetyl-TAGs is a renewable resource with potential industrial uses, including plasticizers, biodegradable lubricants and food emulsifiers.

Research collaborators include Mike Pollard, John Ohlrogge, Jinjie Liu, Adam Rice, Kathleen McGlew and Vincent Shaw with the Great Lakes Bioenergy Research Center at Michigan State University; and Hyunwoo Park and Tom Clemente at the University of Nebraska, Lincoln. The researchers have been supported with a four-year $1.5 million joint US Department of Agriculture and Department of Energy grant.

Resources

  • Jinjie Liu, Henrik Tjellström, Kathleen McGlew, Vincent Shaw, Adam Rice, Jeffrey Simpson, Dylan Kosma, Wei Ma, Weili Yang, Merissa Strawsine, Edgar Cahoon, Timothy P. Durrett, John Ohlrogge (2015a) “Field production, purification and analysis of high-oleic acetyl-triacylglycerols from transgenic Camelina sativa,” Industrial Crops and Products, Volume 65, Pages 259-268 doi: 10.1016/j.indcrop.2014.11.019

  • Liu J, Rice A, McGlew K, Shaw V, Park H, Clemente T, Pollard M, Ohlrogge J, Durrett TP (2015b) “Metabolic engineering of oilseed crops to produce high levels of novel acetyl glyceride oils with reduced viscosity, freezing point and calorific value.” Plant Biotechnol J. 13(6):858-65. doi: 10.1111/pbi.12325

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