Cooperative non-specific DNA binding by octamerizing lambda cI repressors: a site-specific thermodynamic analysis.
Cooperative non-specific DNA binding by octamerizing lambda cI repressors: a site-specific thermodynamic analysis.
复制标题
八聚 lambda cI 阻遏物的协同非特异性 DNA 结合：位点特异性热力学分析。
DOI：
10.1006/jmbi.1998.2056
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发表时间：
1998
影响因子：
5.6
通讯作者：
Ackers,GK
中科院分区：
文献类型：
--
作者：
Pray,TR;Burz,DS;Ackers,GK
二聚化和位点特异性结合之间的关系已被表征为以前的野生型和突变型cI阻遏在正确的运营商（OR）的噬菌体λ DNA。然而，高阶低聚物（四聚体和八聚体），也从这些cI分子形成的作用仍然难以捉摸。在这项研究中，一个明确的相关性已经建立了阻遏物寡聚化和非特异性DNA结合活性。定量DNA酶I足迹滴定技术的改良已被用于评估cI阻遏物寡聚体对非特异性OR侧翼λ DNA的饱和程度。除了一个突变体，只有那些阻遏物能够八聚体被发现表现出非特异性DNA结合活性。使用一维，单价，位点特异性伊辛晶格近似，或更传统的，多价晶格方法准确地模拟非特异性相互作用。有人发现，阻遏物寡聚体的非特异性DNA结合是高度合作的，并在OR位点特异性结合积极独立。此外，非特异性结合的耦合自由能解析为每个阻遏物的位点特异性结合相似，这表明类似的结构元件可以介导这两个结合过程的合作组件。有人提出，阻遏物分子的组装状态调节其对特异性和非特异性DNA序列的相对亲和力。这些特异性是由组装状态信息从C-末端结构域（其介导自缔合和协同性）到N-末端结构域（其主要介导DNA结合）的传输来变构调节的。虽然二聚体对其OR内的同源位点具有高亲和力，但四聚体和八聚体可优先识别非特异性DNA序列。在这项研究中开发的概念和发现可能有助于定量表征的特异性之间的关系，并在其他系统中，利用多种模式的DNA结合协同非特异性结合。
Relationships between dimerization and site-specific binding have been characterized previously for wild-type and mutant cI repressors at the right operator (OR) of bacteriophage lambda DNA. However, the roles of higher-order oligomers (tetramers and octamers) that are also formed from these cI molecules have remained elusive. In this study, a clear correlation has been established between repressor oligomerization and non-specific DNA-binding activity. A modification of the quantitative DNase I footprint titration technique has been used to evaluate the degree of saturation of non-specific, OR-flanking λ DNA by cI repressor oligomers. With the exception of one mutant, only those repressors capable of octamerizing were found to exhibit non-specific DNA-binding activity. The non-specific interaction was accurately modeled using either a one-dimensional, univalent, site-specific Ising lattice approximation, or a more traditional, multivalent lattice approach. It was found that non-specific DNA-binding by repressor oligomers is highly cooperative and energetically independent from site-specific binding at OR. Furthermore, the coupling free energy resolved for non-specific binding was similar to that of site-specific binding for each repressor, suggesting that similar structural elements may mediate the cooperative component of both binding processes. It is proposed that the state of assembly of the repressor molecule modulates its relative affinity for specific and non-specific DNA sequences. These specificities are allosterically regulated by the transmission of assembly-state information from the C-terminal domain, which mediates self-association and cooperativity, to the N-terminal domain, which primarily mediates DNA-binding. While dimers have a high affinity for their cognate sites within OR, tetramers and octamers may preferentially recognize non-specific DNA sequences. The concepts and findings developed in this study may facilitate quantitative characterization of the relationships between specific, and non-specific binding in other systems that utilize multiple modes of DNA-binding cooperativity.