Geranyl diphosphate synthase: Cloning, expression, and characterization of this prenyltransferase as a heterodimer
Geranyl diphosphate synthase: Cloning, expression, and characterization of this prenyltransferase as a heterodimer
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DOI：
10.1073/pnas.96.23.13062
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发表时间：
1999-11-09
影响因子：
11.1
通讯作者：
Croteau, R
中科院分区：
文献类型：
--
作者：
Burke, CC;Wildung, MR;Croteau, R
从留兰香油腺中分离纯化了香叶基二磷酸合成酶，该酶催化二甲基烯丙基二磷酸和异戊烯基二磷酸缩合生成香叶基二磷酸，后者是单萜生物合成的关键前体。从28和37 kDa的纯蛋白质产生的肽片段显示的氨基酸序列匹配的两个cDNA克隆通过随机筛选薄荷油腺cDNA文库。这两种蛋白质的推导序列显示出一些相似性，现有的异戊二烯基转移酶。并且两者都含有质体靶向序列。每个cDNA单独表达没有产生可检测的异戊二烯基转移酶活性，然而，两者共同表达产生功能性香叶基二磷酸合酶。针对每种蛋白质的抗体被用来证明这两个亚基都需要产生催化活性的天然和重组酶，从而证实香叶基二磷酸合酶是异二聚体。
Geranyl diphosphate synthase, which catalyzes the condensation of dimethylallyl diphosphate and isopentenyl diphosphate to geranyl diphosphate, the key precursor of monoterpene biosynthesis, was purified from isolated oil glands of spearmint. Peptide fragments generated from the pure proteins of 28 and 37 kDa revealed amino acid sequences that matched two cDNA clones obtained by random screening of a peppermint-oil gland cDNA library. The deduced sequences of both proteins showed some similarity to existing prenyltransferases. and both contained a plastid-targeting sequence. Expression of each cDNA individually yielded no detectable prenyltransferase activity; however, coexpression of the two together produced functional geranyl diphosphate synthase. Antibodies raised against each protein were used to demonstrate that both subunits were required to produce catalytically active native and recombinant enzymes, thus confirming that geranyl diphosphate synthase is a heterodimer.