GENETIC-EVIDENCE THAT THE XYLS REGULATOR OF THE PSEUDOMONAS TOL META OPERON CONTROLS THE PM PROMOTER THROUGH WEAK DNA-PROTEIN INTERACTIONS
GENETIC-EVIDENCE THAT THE XYLS REGULATOR OF THE PSEUDOMONAS TOL META OPERON CONTROLS THE PM PROMOTER THROUGH WEAK DNA-PROTEIN INTERACTIONS
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DOI：
10.1128/jb.176.11.3171-3176.1994
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发表时间：
1994-06-01
影响因子：
3.2
通讯作者：
DELORENZO, V
中科院分区：
文献类型：
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作者：
KESSLER, B;HERRERO, M;DELORENZO, V
在存在和不存在苯甲酸酯诱导剂的情况下，已在携带 Pm-lacZ 融合体和不同配置的 xylS 基因的特化大肠杆菌菌株中检查了恶臭假单胞菌 TOL 质粒元操纵子的 Pm 启动子在苯甲酸诱导剂存在和不存在的情况下被其同源 XylS 激活蛋白激活，其中所有控制元件都以接近天然条件和化学计量存在。染色体 Pm-xylX::lacZ 融合体的表达主要依赖于在低 xylS 基因剂量下添加效应子，但这种依赖性随着调节子水平的增加而降低，以至于在没有任何芳香效应子的情况下，过量产生的 XylS 可以将表达水平提高到比正常基础水平高 10(4) 倍的水平。在蛋白质固有溶解度允许的细胞内浓度范围内，Pm 活性从未达到规定的饱和水平，因此表明 XylS 对 Om(R) 和 Om(L)（分别为 Om 右半位点和左半位点）操纵子序列的占用程度较低。转录干扰实验证实了这一点，表明活性XylS占据Pm的频率较低。这一特性允许通过细胞内 XylS 浓度的变化来微调体内 Pm 活性，正如当前模型中预测的那样，以解释 TOL 操纵子的协调调节。
The activation of the Pm promoter of the meta operon of the TOL plasmid of Pseudomonas putida by its cognate XylS activator protein in the-presence and absence of benzoate inducers has been examined in specialized Escherichia coli strains carrying Pm-lacZ fusions and the xylS gene in different configurations in which all controlling elements are present in near native conditions and stoichometry. Expression of a chromosomal Pm-xylX::lacZ fusion was primarily dependent on the addition of an effector at a low xylS gene dosage, but such dependency decreased with increasing levels of the regulator, to the point that hyperproduced XylS could, in the absence of any aromatic effector, raise expression to a level 10(4)-fold higher than normal basal levels. Pm activity never reached a defined saturation level within the range of intracellular concentrations permitted by the intrinsic solubility of the protein, thus suggesting a low degree of occupancy of the Om(R) and Om(L) (Om right and left half-sites, respectively) operator sequences by XylS. This was confirmed by transcription interference experiments, which indicated that the frequency of occupation of Pm by active XylS is low. This property permits a fine tuning of Pm activity in vivo through changes in intracellular XylS concentrations, as is predicted in current models to account for the coordinated regulation of TOL operons.