The overall objective of the project is to understand the metabolism and control of metabolism of fatty acids at the molecular level. Studies of the mechanism of fatty acid biosynthesis, its alteration and regulation will be pursued in animal and microbial systems. The fatty acid synthetases from animal tissues and yeast will be prepared and their properties will be studied. The mechanism of fatty acid synthesis and the structure-function relationship between the catalytic domains of the subunit proteins of this multifunctional enzyme are under investigation. The need of the dimer form of the enzyme to affect carbon-carbon bond formation in the beta-ketoacyl synthetase site of the enzyme is being studied. Also, the various component activities of the synthetase will be isolated after controlled proteolysis of the native enzyme. The properties of these individual catalytic domains are being studied. The synthesis of the fatty acid synthetase is under active investigation. The mRNA coding for the synthetase has been isolated from the uropygial gland of goose, lactating rat mammary gland, and insulin induced chick embryo livers. The purified mRNAs translate in cell free ribosomes into proteins which have the same molecular weight as native synthetases and exhibit the same immunochemical properties as the native enzyme. The cDNA is being prepared, inserted into vector plasmid and cloned in appropriate host cells. The clones containing the cDNA plasmid will be identified and utilized in the preparation of sufficient amounts of cDNA that will be used in nucleotide sequence studies and in isolating and characterizing pure synthetase mRNA and genes.
