P-Ketoacyl-Acyl Carrier Protein Synthetase

/3-Ketoacyl-acyl carrier protein (ACP) synthetase catalyzes the condensation reaction of fatty acid synthesis in Escherichia coli. The homogeneous enzyme reacts with hexanoylCoA to form hexanoyl-enzyme which was isolated and characterized. Hexanoyl-enzyme contains 2 mol of hexanoate/mol of enzyme (molecular weight 66,000); it is labile at alkaline pH, and it reacts with neutral hydroxylamine to form hexanoyl hydroxamic acid. Hexanoate was cleaved from the enzyme when hexanoyl-enzyme was subjected to performic acid oxidation. These properties indicate that hexanoylenzyme is a thioester. Studies of the circular dichroism spectra of fully acylated and nonacylated forms of the enzyme indicated that the secondary structure of the enzyme is relatively unperturbed by the presence of the hexanoyl groups. An a helical content of 65% was estimated for the enzyme from the circular dichroism spectrum. Hexanoyl-enzyme is active in both partial reactions that comprise the P-ketoacylACP synthetase reaction; it reacts with ACP to form hexanoyl-ACP and with malonyl-ACP to form fi-ketooctanoylACP. Although the hexanoate of hexanoyl-enzyme is transferred very rapidly to ACP, the physiological acceptor in this reaction, it is also transferred very slowly to CoA, dithiothreitol, and 2-mercaptoethanol, indicating that the enzyme can react nonspecifically with a number of unrelated mercaptans.