Arró Montserrat, Manzano David, Ferrer Albert
Center for Research in Agricultural Genomics (CRAG), CSIC-IRTA-UAB-UB, Campus UAB Bellaterra, Barcelona, Spain.
Methods Mol Biol. 2014;1153:41-53. doi: 10.1007/978-1-4939-0606-2_4.
Farnesyl diphosphate synthase (FPS) catalyzes the sequential head-to-tail condensation of isopentenyl diphosphate (IPP, C5) with dimethylallyl diphosphate (DMAPP, C5) and geranyl diphosphate (GPP, C10) to produce farnesyl diphosphate (FPP, C15). This short-chain prenyl diphosphate constitutes a key branch-point of the isoprenoid biosynthetic pathway from which a variety of bioactive isoprenoids that are vital for normal plant growth and survival are produced. Here we describe a protocol to obtain highly purified preparations of recombinant FPS and a radiochemical assay method for measuring FPS activity in purified enzyme preparations and plant tissue extracts.
法尼基二磷酸合酶(FPS)催化异戊烯基二磷酸(IPP,C5)与二甲基烯丙基二磷酸(DMAPP,C5)以及香叶基二磷酸(GPP,C10)依次进行头对尾缩合反应,生成法尼基二磷酸(FPP,C15)。这种短链异戊二烯基二磷酸构成了类异戊二烯生物合成途径的一个关键分支点,从该分支点可产生多种对植物正常生长和存活至关重要的生物活性类异戊二烯。在此,我们描述了一种获取重组FPS高度纯化制剂的方案,以及一种用于测定纯化酶制剂和植物组织提取物中FPS活性的放射化学测定方法。