Fatty Acid Biosynthesis Redirected to Medium Chains in Transgenic Oilseed Plants

Abstract

Medium-chain fatty acids (FAs), found in storage lipids of certain plants, are an important renewable resource. Seeds of undomesticated California bay accumulate laurate (12:0), and a 12:0-acyl-carrier protein thioesterase (BTE) has been purified from this tissue. Sequencing of BTE enabled the cloning of a complementary DNA coding for a plastid-targeted preprotein. Expression of the complementary DNA in the seeds of Arabidopsis thaliana resulted in BTE activity, and medium chains accumulated at the expense of long-chain (greater than or equal to 16) FAs. Laurate became the most abundant FA species and was deposited in the storage triacylglycerols. These results demonstrate a mechanism for medium-chain FA synthesis in plants.

Isolation and Characterization of Two Safflower Oleoyl-Acyl Carrier Protein Thioesterase cDNA Clones

“…There are several published studies of acyl-ACP thioesterases from a variety of plant species (7,8,12,17,18,24), and recently the purification and cloning of a 12:0-ACP thioesterase from California bay has been reported (6,30). We used as a starting point for our purification the protocol described by McKeon and Stumpf (12), but found that in our laboratory it did not yield the degree of purity reported by those authors.…”

“…This may have been due, among other reasons, to differences in the variety of safflower tissue used for our isolations, or to the variability in the selectivity and elution properties of ACP-Sepharose columns. Therefore, we added a chromato- (30). As shown in Figure 5, there is a high degree of homology (74% identity) between the safflower and Brassica rapa clones, and a lesser, but significant, homology (31% identity) between the safflower 18:1-ACP thioesterase and the 12:0-ACP thioesterase from California bay.…”

“…Sequences of the CNBr peptides from the 34-kD protein were identical to those obtained from the 40-kD protein, and no differences were seen in sequences arising from pool A versus pool B proteins. Based on incomplete cleavage of some of the peptides, their approximate mol wts, and partial homology to a 12:0-ACP thioesterase from California bay (30), the CNBr peptide sequences were aligned with respect to the amino-terminal sequence, SQ830 (Fig. 2).…”

“…Fatty acid biosynthesis in plants is catalyzed by a series of soluble, discrete enzymes (23) that are localized in the chloroplasts of leaves (16) nia bay (Umbellularia californica), which accumulates high levels of lauric acid (12:0) in its seed triacylglycerols. Recently, a 12:0-ACP-specific thioesterase has been isolated from this species and shown to redirect fatty acid synthesis to the production of laurate in transgenic plants (6,18,30). In plants that accumulate the longer-chain fatty acids, the specificity of the acyl-ACP thioesterase may also play a role in determining the final fatty acid composition of the storage oil.…”

Expression of Lauroyl–Acyl Carrier Protein Thioesterase in Brassica napus Seeds Induces Pathways for Both Fatty Acid Oxidation and Biosynthesis and Implies a Set Point for Triacylglycerol Accumulation

“…Seeds of Brassica napus (cv 212 ⁄86) wild-type plants and plants transformed with the plasmid pCGN3831 (cauliflower mosaic virus 35S lauroyl-acyl carrier protein thioesterase [MCTE]; Voelker et al, 1992) or the plasmid pCGN3828 (napin MCTE; Voelker et al, 1996) were provided by Calgene (Davis, CA). The lines used in this study are derived from transformation events 8 and 11 from pCGN3831 and events 18 and 23 to 198 from pCGN3828.…”