Hatada E N, Naumann M, Scheidereit C
Max-Planck-Institut für Molekulare Genetik, Otto-Warburg-Laboratorium, Berline (Dahlem), Germany.
EMBO J. 1993 Jul;12(7):2781-8. doi: 10.1002/j.1460-2075.1993.tb05939.x.
The vertebrate NF-kappa B/c-rel inhibitors MAD-3/I kappa B alpha, I kappa B gamma/pdI and bcl-3 all share a conserved ankyrin repeat domain (ARD) consisting of six complete repeats, a short acidic motif and/or an incomplete seventh repeat. We present here a detailed analysis of the domain in p105/pdI and MAD-3/I kappa B involved in inhibition of DNA binding and in protein interaction with rel factors. We demonstrate that in both cases an acidic region and six ankyrin-like repeats are sufficient and required for protein interaction with the rel factors. However, for p105/pdI to achieve the high affinity needed to suppress DNA binding, an incomplete seventh repeat is required in addition. Both pdI and MAD-3 associate with rel proteins by forming heterotrimeric complexes, as shown by native gel analysis and by cross-linking. Furthermore, we demonstrate that deletion of only three amino acids in the first repeat converts the subunit specificity of the p105 ARD into that of MAD-3/I kappa B. We conclude that functionally the ARD in these molecules has a modular structure, with different subregions determining the specificity for the NF-kappa B subunits p50 and p65.
脊椎动物的NF-κB/c-rel抑制剂MAD-3/IκBα、IκBγ/pdI和bcl-3都共享一个保守的锚蛋白重复结构域（ARD），该结构域由六个完整的重复序列、一个短酸性基序和/或一个不完整的第七个重复序列组成。我们在此对p105/pdI和MAD-3/IκB中参与抑制DNA结合以及与rel因子进行蛋白质相互作用的结构域进行详细分析。我们证明，在这两种情况下，一个酸性区域和六个锚蛋白样重复序列对于与rel因子的蛋白质相互作用既是充分的也是必需的。然而，对于p105/pdI来说，要实现抑制DNA结合所需的高亲和力，则还需要一个不完整的第七个重复序列。如天然凝胶分析和交联所示，pdI和MAD-3都通过形成异源三聚体复合物与rel蛋白结合。此外，我们证明，仅在第一个重复序列中缺失三个氨基酸就会将p105 ARD的亚基特异性转变为MAD-3/IκB的亚基特异性。我们得出结论，从功能上讲，这些分子中的ARD具有模块化结构，不同的亚区域决定了对NF-κB亚基p50和p65的特异性。
