Studies on the mechanism of fatty acid synthesis. I. Preparation and purification of an enzymes system for reconstruction of fatty acid synthesis.

The fatty acid-synthesizing system of Escherichia coli w fractionated by chromatography on diethylaminoethyl cellulose into three different enzyme fractions: Err, ErrI, a E rv. Incubation of Fraction Err with acetyl coenzyme malonyl coenzyme A, reduced triphosphopyridine nucleotide, and acyl carrier protein (ACP) yielded /GhydroxydecanoylACP, ,l3-hydroxylauryl-ACP, and /3-hydroxymyristyl-ACP variable amounts depending on the Err preparations. A dition of Fraction EIII to reaction mixtures of Fraction E yielded mainly palmitic acid, whereas the addition of Frac tion Erv to the same Err reaction mixture yielded mainl cis-vaccenic acid. Therefore, Fractions ErrI and Erv mu have directed the synthesis of saturated and unsaturated fatty acids from fi-hydroxydecanoyl-ACP, respectively. Isolated @-hydroxyacyl-ACPs could be converted to the saturated homologues by incubation with Fraction E and TPNH or could be elongated to palmitic acid by incuba tion with Fraction ErrI, TPNH, malonyl-CoA, and Fraction %I. Incubation of Fraction ErrI or its subfractions wi P-hydroxydecanoyl-CoA yielded the 2-decenoyl derivative Incubation of Fraction Erv or its subfractions with ,L?-hydrox decanoyl-CoA yielded a mixture of cis-3-decenoyl a trans-2-decenoyl derivatives. The relationship between t two dehydrases and the various products formed are d cussed as well as their role in the synthesis of saturated a unsaturated fatty acids.