The yeast fatty acid synthetase is a complex of two multifunctional proteins, alpha and beta, and is active only in the hexamer form alpha(6)beta(6). Electron microscopic studies of this complex led to a model for the synthetase as an ovate structure consisting of an equatorial plate-like structure (the alpha subunit) to which six arches (the beta subunit) are equally distributed on either side. Studies involving the bifunctional reagent 1,3-dibromo-2-propanone have shown that this arrangement is necessary for the beta-ketoacyl synthetase activity, since its active center requires the juxtaposing of an acyl group attached to an active cysteine-SH of one a subunit and a malonyl group attached to a pantetheine-SH of an adjacent a subunit. This conclusion was based in part on the following facts. 1) Iodoacetamide and dibromopropanone inhibit fatty acid synthesis by inhibiting only the beta-ketoacyl synthetase activity; acetyl-coA, but not malonyl-CoA, protected the synthetase against these inhibitors, suggesting that these reagents react with the site of acetyl binding on the enzyme. 2) Dibromopropanone cross-links the a subunits, yielding oligomers of higher molecular weights. The cross-linking was prevented by pretreatment of the synthetase with acetyl-coA, iodoacetamide, or malonyl-CoA, suggesting the involvement of the active cysteine-SH of the ketoacyl synthetase and the pantetheine-SH of the prosthetic group. 3) Cross-linking of the synthetase with C14-labeled dibromopropanone followed by Baeyer-Villiger oxidation and HC1 hydrolysis yielded equal amounts of C14-labeled sulfones of carboxymethylcysteine and carboxymethylcysteamine, identifying the residues involved at the beta-ketoacyl synthetase sites as the active cysteine-SH of one a subunit and the pantetheine-SH of the adjacent a subunit. These observations led us to postulate a mechanism of fatty acid synthesis where an active center involves two complementary halves of two alpha subunits and the arch beta subunit. In an alpha(6)beta(6) structure, therefore, there exist six sites for fatty acid synthesis, all of which function simultaneously.

• PMID: 7021548
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Reference Type

Journal Article | Research Support, Non-U.S. Gov't | Research Support, U.S. Gov't, Non-P.H.S. | Research Support, U.S. Gov't, P.H.S.

Authors

Stoops JK, Wakil SJ

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