Departmento de Biología Molecular y Bioquímica, Facultad de Ciencias, Universidad de Málaga, Instituto Andaluz de Biotecnología, 29071, Málaga, Spain.
Planta. 2010 Nov;232(6):1471-83. doi: 10.1007/s00425-010-1268-9. Epub 2010 Sep 22.
Glutamate decarboxylase (GAD, EC 4.1.1.15) is a key enzyme in the synthesis of γ-aminobutyric acid (GABA) in higher plants. A complete cDNA encoding glutamate decarboxylase (GAD, EC 4.1.1.15) was characterized from Pinus pinaster Ait, and its expression pattern was studied to gain insight into the role of GAD in the differentiation of the vascular system. Pine GAD contained a C-terminal region with conserved residues and a predicted secondary structure similar to the calmodulin (CaM)-binding domains of angiosperm GADs. The enzyme was able to bind to a bovine CaM-agarose column and GAD activity was higher at acidic pH, suggesting that the pine GAD can be regulated in vivo by Ca(2+)/CaM and pH. A polyclonal antiserum was prepared against the pine protein. GAD expression was studied at activity, protein, and mRNA level and was compared with the expression of other genes during the differentiation of the hypocotyl and induction of reaction wood. In seedling organs, GABA levels closely matched GAD expression, with high levels in the root and during lignification of the hypocotyl. GAD expression was also induced in response to the production of compression wood and its expression matched the pattern of other genes involved in ethylene and 2-oxoglutarate synthesis. The results suggest of a role of GAD in hypocotyl and stem development in pine.
谷氨酸脱羧酶(GAD,EC 4.1.1.15)是高等植物中γ-氨基丁酸(GABA)合成的关键酶。我们从 Pinus pinaster Ait 中鉴定了一个完整的编码谷氨酸脱羧酶(GAD,EC 4.1.1.15)的 cDNA,并研究了其表达模式,以深入了解 GAD 在血管系统分化中的作用。松树 GAD 含有一个保守残基的 C 端区域和一个与被子植物 GAD 的钙调蛋白(CaM)结合域相似的预测二级结构。该酶能够与牛 CaM-琼脂糖柱结合,并且在酸性 pH 值下 GAD 活性更高,这表明松树 GAD 可以在体内受到 Ca(2+)/CaM 和 pH 的调节。我们针对松树蛋白制备了多克隆抗血清。我们在活性、蛋白和 mRNA 水平上研究了 GAD 的表达,并将其与 hypocotyl 分化和诱导反应木过程中其他基因的表达进行了比较。在幼苗器官中,GABA 水平与 GAD 表达密切相关,根和 hypocotyl 木质化过程中水平较高。GAD 表达也响应于压缩木材的产生而诱导,其表达与涉及乙烯和 2-氧戊二酸合成的其他基因的表达模式相匹配。结果表明 GAD 在松树的 hypocotyl 和茎发育中发挥作用。