Laboratory of Enzyme Technology, Department of Agricultural Biotechnology, Agricultural University of Athens, 75 Iera Odos Street, 11855 Athens, Greece.
Funct Integr Genomics. 2012 Mar;12(1):157-72. doi: 10.1007/s10142-011-0248-x. Epub 2011 Sep 10.
In the present work, we describe the characterisation of the glutathione transferase (GST) gene family from Agrobacterium tumefaciens C58. A genome survey revealed the presence of eight GST-like proteins in A. tumefaciens (AtuGSTs). Comparison by multiple sequence alignment generated a dendrogram revealing the phylogenetic relationships of AtuGSTs-like proteins. The beta and theta classes identified in other bacterial species are represented by five members in A. tumefaciens C58. In addition, there are three "orphan" sequences that do not fit into any previously recognised GST classes. The eight GST-like genes were cloned, expressed in Escherichia coli and their substrate specificity was determined towards 17 different substrates. The results showed that AtuGSTs catalyse a broad range of reactions, with different members of the family exhibiting quite varied substrate specificity. The 3D structures of AtuGSTs were predicted using molecular modelling. The use of comparative sequence and structural analysis of the AtuGST isoenzymes allowed us to identify local sequence and structural characteristics between different GST isoenzymes and classes. Gene expression profiling was conducted under normal culture conditions as well as under abiotic stress conditions (addition of xenobiotics, osmotic stress and cold and heat shock) to induce and monitor early stress-response mechanisms. The results reveal the constitutive expression of GSTs in A. tumefaciens and a modulation of GST activity after treatments, indicating that AtuGSTs presumably participate in a wide range of functions, many of which are important in counteracting stress conditions. These functions may be relevant to maintaining cellular homeostasis as well as in the direct detoxification of toxic compounds.
在本工作中，我们描述了根癌农杆菌 C58 谷胱甘肽转移酶 (GST) 基因家族的特征。基因组调查显示根癌农杆菌中有 8 种 GST 样蛋白（AtuGSTs）。通过多重序列比对进行比较生成了一个系统发育树，揭示了 AtuGSTs 样蛋白的系统发育关系。在其他细菌物种中鉴定的β和θ类在根癌农杆菌 C58 中有 5 个成员。此外，还有三个“孤儿”序列不属于任何先前确定的 GST 类。八个 GST 样基因被克隆，在大肠杆菌中表达，并确定了它们对 17 种不同底物的底物特异性。结果表明，AtuGSTs 催化广泛的反应，家族中的不同成员表现出相当不同的底物特异性。使用分子建模预测了 AtuGSTs 的 3D 结构。对 AtuGST 同工酶的比较序列和结构分析的使用使我们能够识别不同 GST 同工酶和类之间的局部序列和结构特征。在正常培养条件下以及在非生物胁迫条件下（添加 Xenobiotics、渗透压胁迫和冷热休克）进行基因表达谱分析，以诱导和监测早期应激反应机制。结果表明 GSTs 在根癌农杆菌中组成型表达，并且在处理后 GST 活性发生调节，表明 AtuGSTs 可能参与广泛的功能，其中许多功能在对抗胁迫条件方面很重要。这些功能可能与维持细胞内稳态以及直接解毒有毒化合物有关。
