Identification and Molecular Characterization of the β-Ketoacyl-[Acyl Carrier Protein] Synthase Component of the Arabidopsis Mitochondrial Fatty Acid Synthase*
By R. Yasuno, P. von Wettstein-Knowles, H. Wada. Journal of Biological Chemistry, 2004
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Substrate specificity of condensing enzymes is a predominant factor determining the nature of fatty acyl chains synthesized by type II fatty acid synthase (FAS) enzyme complexes composed of discrete enzymes. The gene (mtKAS) encoding the condensing enzyme, β-ketoacyl-[acyl carrier protein] (ACP) synthase (KAS), constituent of the mitochondrial FAS was cloned from Arabidopsis thaliana, and its product was purified and characterized. The mtKAS cDNA complemented the KAS II defect in the E. coli CY244 strain mutated in both fabB and fabF encoding KAS I and KAS II, respectively, demonstrating its ability to catalyze the condensation reaction in fatty acid synthesis. In vitro assays using extracts of CY244 containing all E. coli FAS components, except that KAS I and II were replaced by mtKAS, gave C4-C18 fatty acids exhibiting a bimodal distribution with peaks at C8 and C14-C16. Previously observed bimodal distributions obtained using mitochondrial extracts appear attributable to the mtKAS enzyme in the extracts. Although the mtKAS sequence is most similar to that of bacterial KAS IIs, sensitivity of mtKAS to the antibiotic cerulenin resembles that of E. coli KAS I. In the first or priming condensation reaction of de novo fatty acid synthesis, purified His-tagged mtKAS efficiently utilized malonyl-ACP, but not acetyl-CoA as primer substrate. Intracellular targeting using green fluorescent protein, Western blot, and deletion analyses identified an N-terminal signal conveying mtKAS into mitochondria. Thus, mtKAS with its broad chain length specificity accomplishes all condensation steps in mitochondrial fatty acid synthesis, whereas in plastids three KAS enzymes are required.
Subfields: Molecular Biology, Pharmacology, Spectroscopy
View Paper: https://www.semanticscholar.org/paper/a7c192c62140c622fe46bf8186beed4aba100021
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Citations in the corpus
- Acyl-Lipid Metabolism (2013, methodology): “The genes for mitochondrial ACP (At2g44620 and At1g65290) and KAS (At2g04540) have been identified in Arabidopsis (Shintani and Ohlrogge, 1994; Yasuno et al., 2004; Meyer et al., 2007), while those of the other components of mitochondrial FAS have not been identified.”