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首页> 外文期刊>Journal of Molecular Biology >CAP, the -45 region, and RNA polymerase: Three partners in transcription initiation at lacP1 in Escherichia coli
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CAP, the -45 region, and RNA polymerase: Three partners in transcription initiation at lacP1 in Escherichia coli

机译：CAP，-45区和RNA聚合酶：大肠杆菌lacP1转录起始的三个伙伴

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摘要

The lac operon of Escherichia coli is positively regulated by the catabolite activator protein (CAP) bound upstream of the -45 region (CAP binding is centered at -61.5; the -45 region extends from -50 to -38). Certain mutations within the -45 region generate sequences that resemble UP elements in base composition and mimic the stimulation by the rrnBP1 UP element, yielding up to 15-fold stimulation in vivo. These -45 region "UP mutants" are compromised in their CAP stimulation. CAP and UP elements do not act in a fully additive manner in vivo at the lac operon. Transcription assays with the wild-type lac promoter and an Ur mutant of Inc indicate that CAP and UP DNA also fail to act in a completely additive manner in vitro. RNA polymerase can stabilize CAP binding to promoter DNA with a -45 region UP element against a heparin challenge. This shows that CAP and the UP DNA do not compete for the a-CTD as a mechanism for their lack of additivity. CAP and UT elements both demonstrate decreased stimulation of transcription as RNA polymerase concentration is increased from 0.05 to 10 nM in in vitro transcription experiments. In addition CAP also stimulates transcription in a manner that does not decrease as RNA polymerase is varied over this concentration range. This invariable stimulation is by two- to threefold and occurs both in vivo and in vitro. It is not dependent upon the alpha-CTD of RNA polymerase and is maintained in the presence of the AR1 CAP mutant HL159. This two- to threefold in invariable CAP stimulation appears to depend on the -45 region sequence as our -45 region mutants demonstrate different responses to HL159 CAP stimulation in vivo. (C) 1998 Academic Press. [References: 30]
机译：大肠杆菌的lac操纵子受到结合在-45区上游的分解代谢活化蛋白（CAP）的正调控（CAP结合集中在-61.5; -45区从-50延伸到-38）。 -45区域内的某些突变产生的序列类似于碱基组成中的UP元件，并模仿rrnBP1 UP元件的刺激，在体内产生多达15倍的刺激。这些-45区“ UP突变体”在其CAP刺激中受损。 CAP和UP元素在lac操纵子体内不能以完全累加的方式起作用。用野生型lac启动子和Inc的Ur突变体进行的转录分析表明，CAP和UP DNA在体外也无法以完全累加的方式起作用。 RNA聚合酶可通过-45区UP元件稳定CAP与启动子DNA的结合，以抵抗肝素攻击。这表明CAP和UP DNA不竞争α-CTD，因为它们缺乏可加性。在体外转录实验中，随着RNA聚合酶浓度从0.05 nM增加到10 nM，CAP和UT元素均显示出减少的转录刺激。另外，CAP也以不随RNA聚合酶在该浓度范围内变化而降低的方式刺激转录。这种不变的刺激是其两倍或三倍，并且在体内和体外均会发生。它不依赖于RNA聚合酶的alpha-CTD，并在存在AR1 CAP突变体HL159的情况下得以维持。恒定CAP刺激的这种2至3倍似乎取决于-45区序列，因为我们的-45区突变体在体内表现出对HL159 CAP刺激的不同反应。 （C）1998年学术出版社。 [参考：30]

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