Binding of synthetic lactose operator DNAs to lactose represessors.
Binding of synthetic lactose operator DNAs to lactose represessors.
复制标题
合成乳糖操纵基因 DNA 与乳糖抑制因子的结合。
DOI：
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发表时间：
1977
影响因子：
11.1
通讯作者：
M. Caruthers
中科院分区：
文献类型：
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作者：
D. Goeddel;D. Yansura;M. Caruthers
用硝酸纤维素膜过滤实验研究了野生型(SQ)和紧结合(QX86)紫胶阻遏蛋白与合成的21和26个碱基对的紫胶操纵子的相互作用。两个操纵子的抑制子结合特性非常相似，表明两者都含有相同的特定抑制子识别位点。抑制物-算符缔合速率常数(k(A))比解离速率常数(k(D))对离子强度的变化更敏感。与以前观察到的以lambdah80dlac为DNA操纵子的影响相比，k(A)和k(D)对离子强度的响应都相对较小。这些结果表明，在自然条件下，Lac抑制物与26个碱基对操纵子序列以外的DNA区域之间存在静电相互作用。SQ抑制子与任何一个操作符的缔合速率常数都高于对扩散限制反应的预测。我们假设抑制子和操作者之间的长程静电引力加速了缔合反应。非操纵子DNA的存在降低了缔合速率常数，当离子强度为0.05M时，这种影响比在0.20M时更为明显。非操纵子DNA对涉及QX86阻遏子的缔合的k(A)值的降低幅度大于带有SQ阻遏子的缔合。这两种类型的阻遏物与合成操作符相互作用的速率常数也不同。SQ阻滞剂的k(A)和k(D)值均高于QX86阻滞剂。然而，当阻滞剂为QX86时，速率常数对离子强度更为敏感。
The nitrocellulose filter assay was used to study the interactions of wild-type (SQ) and tight-binding (QX86) lac repressors with synthetic lac operators 21 and 26 base pairs long. The repressor binding properties of both operators were very similar, indicating that both contain the same specific repressor recognition sites. The repressor-operator association rate constants (k(a)) were more sensitive than dissociation rate constants (k(d)) to changes in ionic strength. The responses of both k(a) and k(d) to ionic strength were relatively small compared to the effects previously observed with lambdah80dlac as operator DNA. These results suggest that under natural conditions there are electrostatic interactions between lac repressor and DNA regions outside of the 26 base pair operator sequence. Association rate constants for SQ repressor with either operator are higher than have been predicted for diffusion-limited reactions. We postulate that long-range electrostatic attractions between repressor and operator accelerate the association reaction. The presence of nonoperator DNA decreased association rate constants, the effect being more noticeable at an ionic strength of 0.05 M than at 0.20 M. Nonoperator DNA reduced k(a) values for associations involving QX86 repressor to a greater extent than for those with SQ repressor. The two types of repressors also had different rate constants for interactions with synthetic operators. The values for k(a) and k(d) were both higher with SQ repressor than with QX86 repressor. However, the rate constants were more sensitive to ionic strength when the repressor used was QX86.