Acyl Carrier Protein

Synthesis of cis-5-β-hydroxy[2-14C]dodecenoyl-acyl carrier protein (ACP), a postulated intermediate in unsaturated fatty acid synthesis in Escherichia coli, was achieved from cis-3-decenoyl-ACP and [2-14C]malonyl-ACP through the combined action of two enzymes, β-ketoacyl-ACP synthetase and β-keto-acyl-ACP reductase. The conversion of cis-5-β-hydroxy[2-14C]dodecenoyl-ACP to cis-5-trans-2-[2-14C]dodecadienoyl-ACP and to cis-5-[2-14C]-dodecenoyl-ACP was shown utilizing a crude fatty acid synthetase preparation of wild type E. coli. Elongation of cis-5-β-hydroxy[2-14C]dodecenoyl-ACP to long chain unsaturated fatty acids was also shown with this fatty acid synthetase preparation. These studies indicate that cis-5-β-hydroxydodecenoyl-ACP, cis-5-trans-2dodecadienoyl-ACP, and cis-5-dodecenoyl-ACP are intermediates in the synthesis of long chain unsaturated fatty acids. Extracts of a class of fatty acid biosynthetic mutants, fab B, which specifically cannot synthesize unsaturated fatty acids although synthesizing saturated fatty acids normally, were examined for all the above activities. No apparent defect was revealed in any step involved in the utilization of cis-5-β-hydroxy[2-14C]dodecenoyl-ACP for long chain unsaturated fatty acid synthesis. Mutant extracts were also found to contain normal amounts of β-ketoacyl-ACP reductase and β-hydroxyacyl-ACP dehydrase, which apparently function in both the saturated and unsaturated fatty acid biosynthetic pathways, and of enoyl-ACP reductase. It is concluded that fab B mutants contain a mutation in an enzyme which catalyzes a reaction unique to the unsaturated fatty acid pathway which is yet unknown.