Chen Y Q, Sengchanthalangsy L L, Hackett A, Ghosh G
Department of Chemistry and Biochemistry, University of California at San Diego, La Jolla, CA 92093-0359, USA.
Structure. 2000 Apr 15;8(4):419-28. doi: 10.1016/s0969-2126(00)00123-4.
The NF-kappaB family of dimeric transcription factors regulates the expression of several genes by binding to a variety of related DNA sequences. One of these dimers, p65(RelA), regulates a subclass of these targets. We have shown previously that p65 binds to the 5'-GGAA T TTTC-3' sequence asymmetrically. In that complex one subunit base specifically interacts with the preferred 5' half site and the other subunit binds non-specifically to the 3' half site.
Here we describe the crystal structures of two new p65-DNA complexes. One complex contains a pseudosymmetric 5'-GGAA T TTCC-3' DNA sequence taken from the enhancer of the gene encoding interleukin 8 (IL-8) and the other contains the asymmetric 5'-GGAA T TCCC-3' target DNA taken from the enhancer of the gene encoding type VII collagen. As expected, the global positioning of the dimer on both DNA targets is roughly symmetric, however, the hydrogen-bonding patterns at the protein-DNA interfaces differ significantly. One of the p65 monomers in complex with the asymmetric DNA binds to an extra base pair located immediately upstream of the 5'-GGAA-3' half site. We also show that p65 binds to these targets with almost equal affinity and that different residues have variable roles in binding different kappaB targets.
Taken together, these structures reveal that p65 exhibits the unique capability to specifically bind DNA targets of variable lengths from four to ten base pairs. Also, the small protein segment Arg41-Ser42-Ala43 is at least partially responsible for flexibility in DNA-binding modes.
二聚体转录因子NF-κB家族通过与多种相关DNA序列结合来调控多个基因的表达。这些二聚体之一，即p65（RelA），调控这些靶标的一个亚类。我们之前已经表明，p65以不对称方式结合到5'-GGAA T TTTC-3'序列上。在该复合物中，一个亚基碱基特异性地与优先的5'半位点相互作用，而另一个亚基非特异性地结合到3'半位点。
在此我们描述了两种新的p65-DNA复合物的晶体结构。一种复合物包含取自编码白细胞介素8（IL-8）基因增强子的假对称5'-GGAA T TTCC-3' DNA序列，另一种包含取自编码VII型胶原基因增强子的不对称5'-GGAA T TCCC-3'靶DNA。正如预期的那样，二聚体在两个DNA靶标上的整体定位大致对称，然而，蛋白质-DNA界面处的氢键模式却有显著差异。与不对称DNA形成复合物的p65单体之一结合到位于5'-GGAA-3'半位点紧邻上游的一个额外碱基对。我们还表明，p65以几乎相等的亲和力结合这些靶标，并且不同的残基在结合不同的κB靶标中具有可变的作用。
综上所述，这些结构揭示p65具有独特的能力，能够特异性结合长度从四到十个碱基对不等的DNA靶标。此外，小蛋白片段Arg41-Ser42-Ala43至少部分负责DNA结合模式的灵活性。
