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首页> 外文期刊>Journal of bacteriology >Regulation of proline utilization in Salmonella typhimurium: a membrane-associated dehydrogenase binds DNA in vitro.
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Regulation of proline utilization in Salmonella typhimurium: a membrane-associated dehydrogenase binds DNA in vitro.

机译：鼠伤寒沙门氏菌中脯氨酸利用的调节：膜相关的脱氢酶在体外结合DNA。

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The PutA protein is a membrane-associated enzyme that catalyzes the degradation of proline to glutamate. Genetic evidence suggests that in the absence of proline, the PutA protein also represses transcription of the putA and putP genes. To directly determine whether PutA protein binds to the put control region, we analyzed gel retardation of put control region DNA by purified PutA protein in vitro. The put control region is 420 bp. Purified PutA protein bound specifically to several nonoverlapping fragments of control region DNA, indicating the presence of multiple binding sites in the control region. Electrophoretic abnormalities and behavior of circularly permuted fragments of control region DNA indicate that it contains a region of intrinsically curved DNA. To determine whether the multiple binding sites or the DNA curvature are important in vivo, two types of deletions were constructed: (i) deletions that removed sequences predicted to contribute to DNA curvature as well as potential operator sites and (ii) deletions that removed only potential operator sites. Both types of deletions increased expression of the put genes but were still induced by proline, indicating that multiple cis elements are involved in repression. These data suggest a model for put repression that invokes the formation of a complex between PutA protein molecules bound at different sites in the control region, brought into proximity by a loop of curved DNA.
机译：PutA蛋白是一种与膜相关的酶，可催化脯氨酸降解为谷氨酸。遗传证据表明，在脯氨酸不存在的情况下，PutA蛋白还会抑制putA和putP基因的转录。为了直接确定PutA蛋白是否结合到put控制区，我们通过体外纯化的PutA蛋白分析了put控制区DNA的凝胶阻滞作用。投放控制区域为420 bp。纯化的PutA蛋白与控制区DNA的几个非重叠片段特异性结合，表明控制区中存在多个结合位点。电泳异常和控制区DNA的圆形排列片段的行为表明，它包含一个固有弯曲的DNA区域。为了确定多个结合位点或DNA曲率在体内是否重要，构建了两种类型的缺失：（i）删除了预计有助于DNA曲率的序列的删除以及潜在的操纵子位点，以及（ii）仅删除了这些删除潜在的运营商网站。两种类型的缺失均增加了put基因的表达，但仍被脯氨酸诱导，表明多种顺式元件参与了抑制。这些数据表明了一种抑制表达的模型，该模型调用了结合在控制区域不同位点的PutA蛋白分子之间复合物的形成，并通过弯曲的DNA环使其接近。

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