High level expression of chorismate pyruvate-lyase (UbiC) and HMG-CoA reductase in hairy root cultures of Lithospermum erythrorhizon.
High level expression of chorismate pyruvate-lyase (UbiC) and HMG-CoA reductase in hairy root cultures of Lithospermum erythrorhizon.
复制标题
DOI：
10.1093/pcp/pcf106
复制
发表时间：
2002-08
影响因子：
4.9
通讯作者：
A. Köhle;S. Sommer;K. Yazaki;A. Ferrer;A. Boronat;Shu‐Ming Li;L. Heide
中科院分区：
文献类型：
--
作者：
A. Köhle;S. Sommer;K. Yazaki;A. Ferrer;A. Boronat;Shu‐Ming Li;L. Heide
紫草素是一种红色的萘醌色素，由紫草属植物紫草（Boraginaceae）的细胞培养物产生。它是由两个关键前体，4-羟基苯甲酸酯（4HB）和香叶酰二磷酸（GPP）生物合成衍生的。细菌ubiC基因编码chorismate pyruate -lyase (CPL)，将choris酸转化为4-羟基苯甲酸盐，该基因在L. erythrorhizon中受强（ocs）(3)ma -启动子控制表达。这引入了一种高效的生物合成途径，即从choris酸一步反应，除了内源性多步苯丙烷途径。用[1,7-(13)C(2)]莽草酸饲喂实验表明，在最活跃的转基因品系中，73%的4HB是通过遗传引入途径合成的。而转基因植株的CPL活性与4HB糖苷和紫草素积累没有相关性。HMG-CoA还原酶（HMGR）参与红藓菌GPP的生物合成。在（ocs）(3)mas启动子的控制下，拟南芥两种HMGR1在紫草中表达。使用完整的酶只获得了适度的酶活性增加，但使用HMGR1的可溶性胞质结构域获得了高活性。即使可溶性HMGR高表达，紫草素的积累也保持不变。
Shikonin, a red naphthoquinone pigment, is produced by cell cultures of Lithospermum erythrorhizon (Boraginaceae). It is biosynthetically derived from two key precursors, 4-hydroxybenzoate (4HB) and geranyldiphosphate (GPP). The bacterial ubiC gene, encoding chorismate pyruvate-lyase (CPL) which converts chorismate to 4-hydroxybenzoate, was expressed in L. erythrorhizon under the control of the strong (ocs)(3)mas-promoter. This introduced an efficient biosynthetic pathway to 4HB, i.e. a one-step reaction from chorismate, in addition to the endogeneous multi-step phenylpropanoid pathway. Feeding experiments with [1,7-(13)C(2)]shikimic acid showed that in the most active transgenic line, 73% of 4HB was synthesized via the genetically introduced pathway. However, there was no correlation between CPL activity and 4HB glucoside or shikonin accumulation in the transgenic lines. HMG-CoA reductase (HMGR) is involved in the biosynthesis of GPP in L. erythrorhizon. Two forms of HMGR1 of Arabidopsis thaliana were expressed in Lithospermum under control of the (ocs)(3)mas promoter. Only moderate increases in enzyme activity were obtained with the complete enzyme, but high activity was achieved using the soluble cytosolic domain of HMGR1. Shikonin accumulation remained unchanged even upon high expression of soluble HMGR.