Ketoacyl-ACP synthetase

After conversion to acetyl-AGP and malonyl-AGP, two carbons of the malonyl-AGP are introduced via the condensing enzyme, 6-ketoacyl-ACP synthetase. Loss of the malonyl carboxyl drives the reaction and in the first step of the sequence acetoacetyl-AGP is formed. The B-ketoacyl-ACP is then reduced to B-hydroxyacyl-AGP by NADPH and the enzyme B-ketoacyl-AGP reductase. The hydroxy acid is dehydrated to form a trans-2.3-enov1-ACP which can be reduced by NADH or NAOPH to the saturated AGP derivative (butyrate in the first series of steps). Gondensation with malony 1-ACP is then repeated and the cycle continues to produce acyl-ACP derivatives with two additional carbon atoms until palmitoy1-ACP results. A second B-ketoacyl-ACP synthetase accomplishes addition of two more malonyl carbon atoms to allow the formation of stearoyl-ACP. The B-ketoacyl-ACP synthetase has been shown to be a separate enzyme since it is more easily inhibited by arsenite and is less sensitive to the antibiotic, cerulenin, than the B-ketoacyl-ACP synthetase forming C to C g keto acids. 

The end product of the Type II fatty acid synthetase of plants is palmitoyl-ACP (Stumpf, 1980) which then serves as the substrate for the elongation system (palmitate elongase). Palmitate elongase has been studied in a number of soluble and membrane-bound subcellular fractions from plants (cf. Harwood, 1979). The enzyme is sensitive to arsenite in contrast to the Type II synthetase which is inhibited by cerulenin. These selective inhibitions seem to be related to the properties of the )8-ketoacyl-ACP synthetase. Two of these enzymes have been purified from spinach leaves (Shimakata and Stumpf, 19826). One will condense a broad range of primer units (up to C14) and is sensitive to cerulenin whereas the second is specific for palmitoyl-ACP and is inhibited by arsenite. The latter enzyme can, therefore, be regarded as a key part of the Type III synthetase, palmitate elongase. 

A ) -ketoacyl-ACP synthetase condenses acetyl ACP and malonyl ACP to form acetoacetyl ACP, CO2, and ACP. The enzyme has been crystallized and has a molecular weight of 66,000. 

Kinney, A.J., Hitz, W.D. and Yadav, N.S. (1990) Stearoyl-ACP desaturase and a P-ketoacyl-ACP synthetase from developing soybean seeds, in Plant Lipid Biochemistry, Structure and Utilization, eds P.J. Quinn and J.L. Harwood, Portland, London, pp. 126-128. 

The most complete examination of the relative rates for the individual activities listed above, including Cuphea, Scifflower and rapeseeds, suggests acetyl-CoA ACP transacylase and B-ketoacyl-ACP synthetase as rate-limiting. ... 

R. W. Hendren and K. Bloch, Fatty acid synthesis from Euglena gracilis. Separation of component activities of the ACP-dependent fatty acid syntehtase and partial purification of the g-ketoacyl-ACP synthetase, 3. Biol. Chem. 255 150 f (1980). 

Partial separation of 6-ketoacyl-ACP synthetase, 6-ketoacyl-ACP reductase, acetyl CoA ACP transacylase and malonyl-CoAiACP transacylase was achieved from barley chloroplasts . From avocado fruit, Caughey and Kekwlck purified the B-ketoacyl-ACP reductase and malonyl-CoA. ACP acyltransferase to homogeneity and also purified the enoyl-ACP reductase. However, the most thorough study was that by Shlmakata and Stumpf mainly with spinach leaves. Purifications of acetyl CoArACP transacylase, B-ketoacyl-ACP synthetase B-ketoacyl-ACP synthetase II, B-ketoacyl-ACP reductase... 

Since chloroplast fatty acid synthetase could associate with thylakoid membranes, especially in lettuce, and because fatty acid synthesis is likely to Integrate with membrane lipid formation in vivo, we tested the effect of different chloroplast fractions on the pattern of products (Table 3). The results showed some differences in the patterns produced and, in particular, that the 0-ketoacyl-ACP synthetase II of pea may be less tightly associated with membranes than synthetase I. 

T. Shimakata and P.K. Stumpf, Purification and characterization of 3-ketoacyl-ACP synthetase 1 from Spinacia oleracea leaves. Arch. Biochem. Biophys. 220 39 (1983). 

Potential sources of differences in fatty acid compostion among different groups of organisms, therefore, are the use of different substrates as the chain extenders (i.e. branched vs. straight-chain compounds) and metabolic control of fatty acid chain termination. These latter are complex and regulated by a variety of nutritional, developmental and environmental factors. Chain termination mechanisms can involve P-ketoacyl-ACP synthetase specificity (from 2-16 carbons) palmitoyl-ACP P-ketostearoyl-ACP synthetase (16-18 carbons) stearoyl-(oleoyl)-CoA P-ketoeicosanoyl-CoA synthetase (18-20 caibons) FAS systems catalyzing condensation of acyl-CoA and malonyl-CoA (20-n carbons) or, for 2-carbon... 

The biosynthesis of fatty acids in plants Is catalysed by a type II, dissociable, fatty acid synthetase (FAS) made up of at least six catalytic polypeptides and a central, acyl carrier protein (ACP). There are three p Ketoacyl-ACP synthetases (KAS) KAS 1, catalysing the initial reaction between acetyl ACP and malonyl ACP KAS 2, catalysing the addition of acetyl ACP to the elongating fatty acid chain and the recently identified KAS 3. A further enzyme implicated in the initiation of FAS is acetyl CoA ACP transacylase (AC AT) [3]. AC AT catalyses the formation of acetyl ACP. KAS 3, however does not utilise acetyl ACP, instead the condensation occurs between malonyl-ACP and acetyl CoA. There has recently been much discussion about the possible roles of KAS 3 and ACAT which appear to have partially overlapping functions KAS 3 activity has been purified to homogeneity from spinach [1] and E. coli [4]. In both the purified enzyme has the ability to load an acetyl group from acetyl CoA, ACAT activity. Purification of ACAT from E. coli has been reported [3], but as this work predates the discovery of KAS 3 it is not clear whether this activity is resolvable from KAS 3. There is still confusion as to whether plants posses distinct ACAT activity since the assays that have been used for both enzymes contain ACP. 

Acetyl-CoA ACP transacylase is a thiolactomycin-sensitive enzyme which catalyzes what has often been regarded as the slowest of all the partial reactions. However, the recent observation that in spinach leaves and all other plant tissues examined there is an acetoacetyl-ACP synthetase which bypasses this reaction (and which has much higher activity) sheds doubt on the physiological importance of acetyl-CoA ACP transacylase. The acetoacetyl-ACP synthetase, sometimes called the short-chain condensing enzyme, is cerulenin insensitive and can, therefore, be differentiated easily from the )8-ketoacyl-ACP synthetase I, which is the main condensing enzyme. The latter has been purified recently to homogeneity for the first time. " ... 

Very recently a third condensing enzyme has been reported in E. coli. This condensing enzyme is distinctly different from the other -ketoacyl-ACP synthetases in E. coli in that it is (a) cerulenin-insensitive (b) specific for very short chain acyl-ACPs and (c) prefers acetyl-CoA over acetyl-ACP. It has been termed acetoacetyl-ACP synthetase. A similar enzyme has... 

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