Seldeen Kenneth L, McDonald Caleb B, Deegan Brian J, Farooq Amjad
Department of Biochemistry & Molecular Biology and the UM/Sylvester Braman Family Breast Cancer Institute, Leonard Miller School of Medicine, University of Miami, 1011 NW 15th Street, Gautier Building, Room 214, Miami, FL 33136, USA.
Arch Biochem Biophys. 2008 May 1;473(1):48-60. doi: 10.1016/j.abb.2008.02.024. Epub 2008 Feb 26.
In response to mitogenic stimuli, the heterodimeric transcription factor Jun-Fos binds to the promoters of a diverse array of genes involved in critical cellular responses such as cell growth and proliferation, cell cycle regulation, embryogenic development and cancer. In so doing, Jun-Fos heterodimer regulates gene expression central to physiology and pathology of the cell in a specific and timely manner. Here, using the technique of isothermal titration calorimetry (ITC), we report detailed thermodynamics of the bZIP domains of Jun-Fos heterodimer to synthetic dsDNA oligos containing the TRE and CRE consensus promoter elements. Our data suggest that binding of the bZIP domains to both TRE and CRE is under enthalpic control and accompanied by entropic penalty at physiological temperatures. Although the bZIP domains bind to both TRE and CRE with very similar affinities, the enthalpic contributions to the free energy of binding to CRE are more favorable than TRE, while the entropic penalty to the free energy of binding to TRE is smaller than CRE. Despite such differences in their thermodynamic signatures, enthalpy and entropy of binding of the bZIP domains to both TRE and CRE are highly temperature-dependent and largely compensate each other resulting in negligible effect of temperature on the free energy of binding. From the plot of enthalpy change versus temperature, the magnitude of heat capacity change determined is much larger than that expected from the direct association of bZIP domains with DNA. This observation is interpreted to suggest that the basic regions in the bZIP domains are largely unstructured in the absence of DNA and only become structured upon interaction with DNA in a coupled folding and binding manner. Our new findings are rationalized in the context of 3D structural models of bZIP domains of Jun-Fos heterodimer in complex with dsDNA oligos containing the TRE and CRE consensus sequences. Taken together, our study demonstrates that enthalpy is the major driving force for a key protein-DNA interaction pertinent to cellular signaling and that protein-DNA interactions with similar binding affinities may be accompanied by differential thermodynamic signatures. Our data corroborate the notion that the DNA-induced protein structural changes are a general feature of the bZIP family of transcription factors.
在有丝分裂原刺激下，异二聚体转录因子Jun-Fos与一系列参与关键细胞反应（如细胞生长和增殖、细胞周期调控、胚胎发育和癌症）的基因启动子结合。通过这种方式，Jun-Fos异二聚体以特定且及时的方式调节对细胞生理和病理至关重要的基因表达。在此，我们使用等温滴定量热法（ITC）技术，报告了Jun-Fos异二聚体的bZIP结构域与含有TRE和CRE共有启动子元件的合成双链DNA寡核苷酸的详细热力学情况。我们的数据表明，bZIP结构域与TRE和CRE的结合受焓控制，并且在生理温度下伴随着熵罚。尽管bZIP结构域与TRE和CRE的结合亲和力非常相似，但与CRE结合的自由能的焓贡献比TRE更有利，而与TRE结合的自由能的熵罚比CRE小。尽管它们的热力学特征存在这些差异，但bZIP结构域与TRE和CRE结合的焓和熵都高度依赖温度，并且在很大程度上相互补偿，导致温度对结合自由能的影响可忽略不计。从焓变与温度的关系图来看，确定的热容变化幅度远大于bZIP结构域与DNA直接结合所预期的幅度。这一观察结果被解释为表明bZIP结构域中的碱性区域在没有DNA时基本上是无结构的，只有在与DNA相互作用时以耦合折叠和结合的方式才会变得有结构。我们的新发现结合Jun-Fos异二聚体的bZIP结构域与含有TRE和CRE共有序列的双链DNA寡核苷酸的三维结构模型进行了合理化解释。综上所述，我们的研究表明焓是与细胞信号传导相关的关键蛋白质 - DNA相互作用的主要驱动力，并且具有相似结合亲和力的蛋白质 - DNA相互作用可能伴随着不同的热力学特征。我们的数据证实了DNA诱导的蛋白质结构变化是bZIP转录因子家族的一个普遍特征这一观点。
