LAC REPRESSOR-OPERATOR INTERACTION .1. EQUILIBRIUM STUDIES
LAC REPRESSOR-OPERATOR INTERACTION .1. EQUILIBRIUM STUDIES
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DOI：
10.1016/0022-2836(70)90219-6
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发表时间：
1970-01-01
影响因子：
5.6
通讯作者：
BOURGEOIS, S
中科院分区：
文献类型：
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作者：
RIGGS, AD;SUZUKI, H;BOURGEOIS, S
膜过滤技术detectinglacrepressor-operator复合物进行了讨论，它表明，这种方法检测复合物存在于溶液中过滤前。该方法灵敏（10 - 16摩尔）、准确，适合物理化学实验。我们的结果是一致的想法，只有一个阻遏物结合每个运营商。我们报告了平衡常数（1 × 10− 13 min，我们的标准缓冲液）和几种反应条件对该参数的影响。特别地，结合对离子强度非常敏感，平衡常数的对数与离子强度的平方根成比例。阻遏物亚基和诱导剂（异丙基硫代半乳糖苷）结合活性相比，DNA结合活性的优先损失的证据。描述了测定能够结合DNA的阻遏物浓度的方法。放线菌素D抑制结合，表明至少有一个鸟嘌呤中或附近的operator.We还报告了一个敏感的竞争分析未标记的operator和显示，阻遏不结合变性的DNA，但将结合后的DNA复性。不含半乳糖区的天然大肠杆菌DNA即使在超过300倍重量（10 μg/ml）时也不竞争。采用了因此，与操作符的绑定是极其特定的。
A membrane filter technique for detectinglacrepressor-operator complexes is discussed and it is shown that this method assays for complexes present in solution prior to filtering. The method is sensitive (10−16mole), accurate, and suitable for physical chemical experiments. Our results are consistent with the idea that only one repressor binds per operator. We report on the equilibrium constant (1 × 10−13min our standard buffer) and the effect of several reaction conditions on this parameter. In particular, the binding is very sensitive to ionic strength, the logarithm of the equilibrium constant being proportional to the square root of the ionic strength. Evidence for repressor subunits and a preferential loss of DNA binding activity compared to inducer (isopropyl-thiogalactoside) binding activity is presented. Methods for determining the concentration of repressor capable of binding to DNA are described. Actinomycin D inhibits the binding, suggesting that there is at least one guanine in or near the operator.We also report on a sensitive competition assay for unlabeled operator and show that the repressor does not bind to denatured DNA but will bind following renaturation of the DNA. NativeEscherichia coliDNA without thelacregion does not compete even when a 300-fold weight excess (10 μg/ml.) is used. The binding to the operator is thus extremely specific.