Structure-based analysis of HU-DNA binding.
Structure-based analysis of HU-DNA binding.
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基于结构的 HU-DNA 结合分析。
DOI：
10.1016/j.jmb.2006.10.024
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发表时间：
2007
影响因子：
5.6
通讯作者：
Rice,PhoebeA
中科院分区：
文献类型：
--
作者：
Swinger,KerrenK;Rice,PhoebeA
HU和IHF是原核蛋白，可以在DNA中引起非常大的弯曲。它们以高浓度的形式存在于细菌的类核中，有助于染色体的紧凑。它们还在许多过程中发挥调节辅助因子的作用，如位点特异性重组和复制和转录的启动。HU和IHF已经成为理解DNA弯曲和间接读出序列的范例。虽然IHF显示出显著的序列特异性，但HU优先与某些受损或扭曲的DNA结合。然而，先前在体外研究的结构多样化的HU底物没有一个与最近发表的Anabaena Hu(Ahu)-DNA共晶结构中的扭曲底物完全相同。在这里，我们报告了AHU与共晶结构中的DNA的结合亲和力。这些AHU-DNA络合物的结合自由能范围为∼10-14.5千卡/摩尔，代表纳摩尔到低皮摩尔范围内的Kd值，相对于未扭曲的非特异性∼络合物，最大稳定度至少为6.3千卡/摩尔。我们研究了IHF结合，发现适当的结构扭曲可以极大地提高其亲和力。在结构和相关结合数据耦合的基础上，我们估计了IHF介导的DNA扭结中被缺口(至少0.76千卡/摩尔)解除的构象应变量，并精确地定位了应变的位置。我们发现Ahu在其DNA结合位点的中心有一个富含A+T的区域的序列偏好，与一个异常狭窄的小沟相关。这类似于真核细胞核小体所显示的序列偏好。
HU and IHF are prokaryotic proteins that induce very large bends in DNA. They are present in high concentrations in the bacterial nucleoid and aid in chromosomal compaction. They also function as regulatory cofactors in many processes, such as site-specific recombination and the initiation of replication and transcription. HU and IHF have become paradigms for understanding DNA bending and indirect readout of sequence. While IHF shows significant sequence specificity, HU binds preferentially to certain damaged or distorted DNAs. However, none of the structurally diverse HU substrates previously studied in vitro is identical with the distorted substrates in the recently published Anabaena HU(AHU)–DNA cocrystal structures. Here, we report binding affinities for AHU and the DNA in the cocrystal structures. The binding free energies for formation of these AHU–DNA complexes range from ∼10–14.5 kcal/mol, representing Kdvalues in the nanomolar to low picomolar range, and a maximum stabilization of at least ∼6.3 kcal/mol relative to complexes with undistorted, non-specific DNA. We investigated IHF binding and found that appropriate structural distortions can greatly enhance its affinity. On the basis of the coupling of structural and relevant binding data, we estimate the amount of conformational strain in an IHF-mediated DNA kink that is relieved by a nick (at least 0.76 kcal/mol) and pinpoint the location of the strain. We show that AHU has a sequence preference for an A+T-rich region in the center of its DNA-binding site, correlating with an unusually narrow minor groove. This is similar to sequence preferences shown by the eukaryotic nucleosome.