Proton-linked contributions to site-specific interactions of lambda cI repressor and OR.
Proton-linked contributions to site-specific interactions of lambda cI repressor and OR.
复制标题
质子连接对 lambda cI 阻遏物和 OR 的位点特异性相互作用的贡献。
DOI：
10.1021/bi00480a004
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发表时间：
1990
期刊：
影响因子：
2.9
通讯作者：
Ackers,GK
中科院分区：
文献类型：
--
作者：
Senear,DF;Ackers,GK
用DNase I足迹滴定方法研究了质子活度对CL抑制物与操纵子OR位点特异性相互作用的影响。获得了阻遏子与野生型0R和突变操纵子的每个位点的结合等温线，其中取消了与某些位点的结合。在每个pH(范围5-8)下测定了(在每个操作者构型中)结合和协同作用的吉布斯能。质子关联效应显然解释了三个操作点亲和力差异的很大一部分原因。对抑制物-操纵子结合作用最显著的影响是在酸性pH，因此不涉及已知的与DNA接触的抑制物N末端臂中的碱性基团。此外，质子连接效应在三个操作点上是不同的，由显著不同的导数关系表明，在An=AVH中，d在k/d中。这些结果表明，可能不与DNA接触的可电离阻遏基团和三个阻遏-操纵子位点复合体之间的构象差异是位点特异性机制的重要组成部分。这些研究产生的大量数据库也被用来重新评估该系统中用于描述合作性的传统模型。结果证实，在所有条件下，0R1和OR3之间没有显著的协同作用。然而，某些实验条件的数据显然不符合选择规则，即Or2和Or3之间的合作作用是通过在0R1[Johnson，A.D.，Meyer，BJ，&Ptashne，M.(1979)Proc.娜塔莉。阿卡德。SCI。美国76,5061-5065]，
The effects of proton activity on the site-specific interactions of cl repressors with operator sites 0R were studied by using DNase I footprint titration. Individual-site binding isotherms were obtained for the binding of repressor to each site of wild-type0R and of mutant operators in which binding to some sites is eliminated. The Gibbs energies for binding and for cooperativity (in every operator configuration) were determined at each pH (range 5-8). The proton-linked effects clearly account for a significant fraction of the difference in affinities for thethree operator sites. The most dramatic effects on the repressor-operator binding interactions are at acid pH, and therefore do not involve the basic groups in the repressor N-terminal arm known to contact the DNA. Also, the proton-linked effects are different at the three operator sites as indicated by significantly different derivative relationships, d In k/d In an= AvH. These results implicate ionizable repressor groups which may not contact the DNA and conformational differencesbetween the three repressor-operator site complexes as being important components to the mechanism of site specificity. The extensive data base generated by these studies was also used to reevaluate the traditional models used to describe cooperativity in this system. The results confirm the lack of significant cooperative interaction between 0R1 and Or3 at all conditions. However, the data for some experimental conditions are clearly inconsistent with the (selection) rule, that cooperative interaction between Or2 and Or3 is eliminated by ligation at 0R1 [Johnson, A. D., Meyer, BJ, & Ptashne, M.(1979) Proc. Natl. Acad. Sci. USA 76, 5061-5065],