Fatty acid modifying enzymes from developing seeds of Vernonia galamenensis

#This invention is in the field of plant molecular biology. More specifically, this invention pertains to nucleic acid fragments encoding enzymes involved in fatty acid biosynthesis and modification in plants and seeds.

#Fatty acids bearing chemical modifications in addition to the common double bonds are found in the storage lipids of many oilseeds (Harwood, J. L. (1980) In The Biochemistry of Plants, T. S. Moore Jr., ed. CRC Press, New York, pp 91-116). Some of these modifications functionalize the fatty acid to produce products that are useful in industrial applications; this is opposed to the more common usage of plant-derived lipids as foods. Examples are the use of the hydroxylated fatty acid ricinoleic acid in lubricants, and the short- or medium-carbon chain length fatty acids from palm oil in detergents. In some cases, fatty acid composition of the storage lipids of oilseeds produced in temperate climates can be modified by the addition of genes from exotic sources so that large amounts of unique fatty acids are produced (Ohlrogge, J B. (1994) Plant Physiol. 104, 821-826).

#Epoxidation is among the known modifications to storage lipid fatty acids. The 18-carbon fatty acid 9,10-ene-12,13-epoxide comprises as much as 60% of the total seed fatty acid in species such as Vernonia galamenensis and Euphorbia lagascea (Bafor, M. et al. (1993) Arch. Biochem Biophys 303:145-151). Fatty acids carrying the epoxide modification may find use as plasticizers, in crosslinking coatings applications, and in setting printing inks.

#The invention can be more fully understood from the following detailed description and the accompanying drawing and sequence descriptions which form a part of this application.

#FIG. 1 shows a comparison of the amino acid sequences of the instant Vernonia galamenensis fatty acid desaturase (vs1.05h08), the instant Vernonia galamenensis fatty acid epoxidase (vs1.02c07), a soybean fatty acid desaturase (soy) and a castor bean fatty acid hydroxylase (castor). The eight conserved histidine residues found in all membrane bound fatty acid modifying enzymes of this class are boxed and stippled. An arginine residue found in the fatty acid hydroxylase from castor bean and in the instant Vernonia galamenensis fatty acid epoxidase but not in the fatty acid desaturating enzymes of this class is also boxed and stippled. Other residues in the Vernonia fatty acid epoxidase sequence that are unique to that sequence but conserved in most or all of the other sequences are boxed. Not all amino acid changes are marked: only those in which the change in the epoxidase sequence occurs in a highly conserved region and in which the amino acid present in the epoxidase is different in physical character from the conserved residues in the other sequences are marked.

#The following sequence descriptions and sequence listings attached hereto comply with the rules governing nucleotide and/or amino acid sequence disclosures in patent applications as set forth in 37 C.F.R. .sctn.1.821-1.825.

#This invention relates to isolated nucleic acid fragments encoding all or a substantial portion of fatty acid epoxidizing and desaturase enzymes normally expressed in developing seeds of Vernonia galamenensis that are similar in sequence to other plant, membrane-bound fatty acid desaturases. The invention also relates to the construction of a chimeric gene comprising an nucleic acid fragment encoding all or a portion of the Vernonia galamenensis fatty acid epoxidizing or desaturase enzyme, operably linked in sense or antisense orientation to suitable regulatory sequences, wherein expression of the chimeric gene results in production of altered levels of the desired enzyme in a transformed host cell. The invention also relates to methods of using isolated nucleic acid fragments encoding all or a substantial portion of the fatty acid epoxidizing and desaturase enzymes normally expressed in developing seeds of Vernonia galamenensis.

#In the context of this disclosure, a number of terms shall be utilized. As used herein, an "isolated nucleic acid fragment" is a polymer of RNA or DNA that is single- or double-stranded, optionally containing synthetic, non-natural or altered nucleotide bases. An isolated nucleic acid fragment in the form of a polymer of DNA may be comprised of one or more segments of cDNA, genomic DNA or synthetic DNA.

#The term "Vernonia galamenensis fatty acid modifying enzyme" refers collectively to the Vernonia galamenensis fatty acid epoxidizing enzyme and the Vernonia galamenensis fatty acid desaturase enzyme disclosed in the instant specification.