Prisic Sladjana, Peters Reuben J
Department of Biochemistry, Biophysics, and Molecular Biology, Iowa State University, Ames, IA 50011, USA.
Plant Physiol. 2007 May;144(1):445-54. doi: 10.1104/pp.106.095208. Epub 2007 Mar 23.
Gibberellins (GAs) or gibberellic acids are ubiquitous diterpenoid phytohormones required for many aspects of plant growth and development, including repression of photosynthetic pigment production (i.e. deetiolation) in the absence of light. The committed step in GA biosynthesis is catalyzed in plastids by ent-copalyl diphosphate synthase (CPS), whose substrate, (E,E,E,)-geranylgeranyl diphosphate (GGPP), is also a direct precursor of carotenoids and the phytol side chain of chlorophyll. Accordingly, during deetiolation, GA production is repressed, whereas flux toward these photosynthetic pigments through their common GGPP precursor is dramatically increased. How this is accomplished has been unclear because no mechanism for regulation of CPS activity has been reported. We present here kinetic analysis of recombinant pseudomature CPS from Arabidopsis (Arabidopsis thaliana; rAtCPS) demonstrating that Mg(2+) and GGPP exert synergistic substrate inhibition effects on CPS activity. These results suggest that GA metabolism may be limited by feed-forward inhibition of CPS; in particular, the effect of Mg(2+) because light induces increases in plastid Mg(2+) levels over a similar range as that observed here to affect rAtCPS activity. Notably, this effect is most pronounced in the GA-specific AtCPS because the corresponding activity of the resin acid biosynthetic enzyme abietadiene synthase is 100-fold less sensitive to [Mg(2+)]. Furthermore, Mg(2+) allosterically activates the plant porphobilinogen synthase involved in chlorophyll production. Hence, Mg(2+) may have a broad role in regulating plastidial metabolic flux during deetiolation. Finally, the observed synergistic substrate/feed-forward inhibition of CPS also seems to provide a novel example of direct regulation of enzymatic activity in hormone biosynthesis.
赤霉素（GAs）或赤霉酸是普遍存在的二萜类植物激素，在植物生长和发育的许多方面都发挥着作用，包括在无光条件下抑制光合色素的产生（即去黄化）。赤霉素生物合成的关键步骤由质体中的内根-贝壳杉烯二磷酸合酶（CPS）催化，其底物（E,E,E）-香叶基香叶基二磷酸（GGPP）也是类胡萝卜素和叶绿素植醇侧链的直接前体。因此，在去黄化过程中，赤霉素的产生受到抑制，而通过它们共同的GGPP前体流向这些光合色素的通量则显著增加。由于尚未报道调节CPS活性的机制，所以这一过程是如何实现的尚不清楚。我们在此展示了对拟南芥重组假成熟CPS（rAtCPS）的动力学分析，结果表明Mg(2+)和GGPP对CPS活性具有协同底物抑制作用。这些结果表明，赤霉素代谢可能受到CPS前馈抑制的限制；特别是Mg(2+)的作用，因为光照会使质体中Mg(2+)水平升高，升高范围与本文观察到的影响rAtCPS活性的范围相似。值得注意的是，这种效应在赤霉素特异性的AtCPS中最为明显，因为树脂酸生物合成酶枞酸二烯合酶的相应活性对[Mg(2+)]的敏感性低100倍。此外，Mg(2+)能变构激活参与叶绿素生成的植物胆色素原合酶。因此，Mg(2+)可能在去黄化过程中调节质体代谢通量方面发挥广泛作用。最后，观察到的CPS协同底物/前馈抑制似乎也为激素生物合成中酶活性的直接调节提供了一个新例子。
