The Condensation Reaction of Fatty Acid Synthesis.iii. Identification of the Protein-bound Product of the Reaction and Its Conversion to Long Chain Fatty Acids.

Long chain fatty acid biosynthesis proceeds through a condensation between acetyl coenzyme A and malonyl coenzyme A (1,2) as has now been shown in fatty acid synthetase systems from animal (3), plant (4), and microbial (5-7, Sl) sources. With studies on extracts of Escherichia coli, in which the major product is vaccenic acid, Lennarz, Light, and Bloch (7) have shown and we have confirmed (8) that this mechanism applies to the synthesis of an unsaturated fatty acid as well as to saturated fatty acids. In all these systems, acetyl-CoA forms the 2 carbons at the methyl-terminal end of the fatty acid, whereas the rest of the carbon chain is formed from 2 carbon units derived from malonyl-CoA. A condensation with concomitant decarboxylation has been proposed as the mechanism of fatty acid elongation by malonylCoA. Such a proposal is supported by two types of evidence. One is the isolation of enzyme-bound acetoacetate as the product of a condensation between acetyl-CoA and malonyl-CoA in a purified synthetase system from yeast (5). Such a product is consistent with a condensation-decarboxylation of malonyl-CoA with acetyl-CoA. However, in the isolation of the condensed product, the binding protein was inactivated; therefore, proof was lacking that this condensed product was an intermediate in the synthesis of long chain fatty acids. A second type of evidence linking the condensation-decarboxylation of malonyl-CoA with long chain fatty acid synthesis is the association of the enzymes for the malonyl-CoA-CO2 exchange reaction or the analogous acyl-CoA-dependent decarboxylation of malonyl-CoA with the enzymes of fatty acid synthesis in a number of purified systems (3, 5, 9-11). Furthermore, both the malonyl-CoA-CO2 exchange reaction and over-all fatty acid synthesis are dependent on the same heat-stable and heat-labile protein fractions in fatty acid-synthesizing systems from Cbstridium kluyveri and Escherichia coli (6,8). It is the purpose of this paper to present evidence that acetoacetate formed by the condensation of malonyl-CoA and acetylCoA is bound to the heat-stable protein, Enzyme II. In the presence of acetyl-CoA and malonyl-1 , 314C-CoA, the reaction is visualized as follows.