Rodríguez-Rodríguez Manuel Fernando, Salas Joaquín J, Garcés Rafael, Martínez-Force Enrique
Instituto de la Grasa, CSIC, Avda Padre García Tejero 4, 41012 Seville, Spain.
Instituto de la Grasa, CSIC, Avda Padre García Tejero 4, 41012 Seville, Spain.
Phytochemistry. 2014 Nov;107:7-15. doi: 10.1016/j.phytochem.2014.08.014. Epub 2014 Sep 8.
Acyl-acyl carrier protein (ACP) thioesterases are intraplastidial enzymes that terminate de novo fatty acid biosynthesis in the plastids of higher plants by hydrolyzing the thioester bond between ACP and the fatty acid synthesized. Free fatty acids are then esterified with coenzyme A prior to being incorporated into the glycerolipids synthesized through the eukaryotic pathway. Acyl-ACP thioesterases belong to the TE14 family of thioester-active enzymes and can be classified as FatAs and FatBs, which differ in their amino acid sequence and substrate specificity. Here, the FatA and FatB thioesterases from Camelina sativa seeds, a crop of interest in plant biotechnology, were cloned, sequenced and characterized. The mature proteins encoded by these genes were characterized biochemically after they were heterologously expressed in Escherichia coli and purified. C. sativa contained three different alleles of both the FatA and FatB genes. These genes were expressed most strongly in expanding tissues in which lipids are very actively synthesized, such as developing seed endosperm. The CsFatA enzyme displayed high catalytic efficiency on oleoyl-ACP and CsFatB acted efficiently on palmitoyl-ACP. The contribution of these two enzymes to the synthesis of C. sativa oil was discussed in the light of these results.
酰基 - 酰基载体蛋白（ACP）硫酯酶是质体内的酶，通过水解ACP与合成的脂肪酸之间的硫酯键，终止高等植物质体中的从头脂肪酸生物合成。然后，游离脂肪酸在通过真核途径合成甘油脂之前，先与辅酶A酯化。酰基 - ACP硫酯酶属于硫酯活性酶的TE14家族，可分为FatA和FatB，它们在氨基酸序列和底物特异性上有所不同。在此，对来自荠蓝种子（一种植物生物技术领域备受关注的作物）的FatA和FatB硫酯酶进行了克隆、测序和表征。这些基因编码的成熟蛋白在大肠杆菌中异源表达并纯化后，进行了生化特性分析。荠蓝含有FatA和FatB基因的三种不同等位基因。这些基因在脂质合成非常活跃的扩展组织中表达最强，如发育中的种子胚乳。CsFatA酶对油酰 - ACP显示出高催化效率，而CsFatB对棕榈酰 - ACP作用高效。根据这些结果讨论了这两种酶对荠蓝油脂合成的贡献。
