Functional characterization of the arabidopsis' γ-glutamyltransferase gene family.

γ-谷氨酰转移酶（γ-Glutamyltransferase，GGT）催化谷胱甘肽（glutathione，GSH）催化的第一个关键步骤是谷胱甘肽S-结合物（glutathione S-conjugate，GSX）的降解和半胱氨酸（cysteine，Cys）的再循环。然而，关于GGT的功能，特别是关于该酶在植物中的体内作用，仍有许多方面不清楚。在拟南芥基因组中，有三个编码GGT的基因（AtGGT 1、AtGGT 2、AtGGT 3）。RT-PCR基因表达分析表明AtGGT 1和AtGGT 3组成型表达。相反，AtGGT 2主要在长角果中表达。通过离心分析GGT活性的分布，表明拟南芥具有可溶性和结合的GGT活性。结合GGT未定位于微粒体组分。此外，结合的GGT被含有500 mM NaCl的高离子强度缓冲液溶解，并且在原生质体中未检测到，这一事实表明结合的GGT可能定位于细胞壁。GGT活性分析表明AtGGT 1编码结合GGT，而AtGGT 2和AtGGT 3编码可溶性GGT。为了研究GGT蛋白在拟南芥中的细胞定位，将AtGGT 1、AtGGT 2、AtGGT 3 cDNA与GFP融合，并在相应启动子的调控下表达。

γ-Glutamyltransferase (GGT) has been proposed to catalyze the first committed step in glutathione (GSH) catabolism, degradation of glutathione S-conjugates (GSX) and cysteine (Cys) recycling. However, there are still many aspects about the GGT function, especially concerning the in vivo role of the enzyme in plants, that are not clear. In the Arabidopsis genome there are three genes (AtGGT1, AtGGT2, AtGGT3) coding for GGT. RT-PCR gene expression analysis suggested that AtGGT1 and AtGGT3 are expressed constitutively. In contrast, AtGGT2 is mainly expressed in siliques. Analysis of the distribution of GGT activity by centrifugation indicated that Arabidopsis has soluble and bound GGT activities. Bound GGT was not localized to the microsomal fraction. Besides, the fact that bound GGT was solubilized by high ionic strength buffer containing 500 mM NaCl, and that it was not detected in protoplasts, indicated that bound GGT is likely localized to the cell wall. GGT activity analysis indicated that AtGGT1 codes for a bound GGT, whereas AtGGT2 and AtGGT3 code for soluble GGTs. In order to carefully analysis the intracellular localization of GGT proteins, AtGGT1, AtGGT2, AtGGT3 cDNAs fused to GFP and expressed under the control of their corresponding promoter were introduced in Arabidopsis.