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首页> 外文期刊>Journal of Molecular Biology >Dimerisation of the UBA domain of p62 inhibits ubiquitin binding and regulates NF-kappaB signalling.
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Dimerisation of the UBA domain of p62 inhibits ubiquitin binding and regulates NF-kappaB signalling.

机译：p62的UBA结构域的二聚化可抑制泛素结合并调节NF-κB信号传导。

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摘要

The ubiquitin (Ub)-binding p62 scaffold protein (encoded by the SQSTM1 gene) regulates a diverse range of signalling pathways leading to activation of the nuclear factor kappa B (NF-kappaB) family of transcription factors and is an important regulator of macroautophagy. Mutations within the gene encoding p62 are commonly found in patients with Paget's disease of bone and largely cluster within the C-terminal ubiquitin-associated (UBA) domain, impairing its ability to bind Ub, resulting in dysregulated NF-kappaB signalling. However, precisely how Ub-binding is regulated at the molecular level is unclear. NMR relaxation dispersion experiments, coupled with concentration-dependent NMR, CD, isothermal titration calorimetry and fluorescence kinetic measurements, reveal that the p62 UBA domain forms a highly stable dimer (K(dim) approximately 4-12 microM at 298 K). NMR analysis shows that the dimer interface partially occludes the Ub-binding surface, particularly at the C-terminus of helix 3, making UBA dimerisation and Ub-binding mutually exclusive processes. Somewhat unusually, the monomeric UBA appears to be the biologically active form and the dimer appears to be the inactive one. Engineered point mutations in loop 1 (E409K and G410K) are shown to destabilise the dimer interface, lead to a higher proportion of the bound monomer and, in NF-kappaB luciferase reporter assays, are associated with reduced NF-kappaB activity compared with wt-p62.
机译：泛素（Ub）结合的p62支架蛋白（由SQSTM1基因编码）调节多种信号通路，导致转录因子核因子κB（NF-κB）家族的激活，并且是巨噬细胞自噬的重要调节剂。编码p62的基因内的突变通常在患有骨Paget病的患者中发现，并且在C端泛素相关（UBA）域内大量聚集，从而削弱了其与Ub结合的能力，从而导致NF-κB信号传导失调。然而，不清楚如何在分子水平上调节Ub结合。 NMR弛豫分散实验，再加上浓度依赖的NMR，CD，等温滴定热法和荧光动力学测量，揭示p62 UBA域形成了高度稳定的二聚体（在298 K处K（dim）约为4-12 microM）。 NMR分析表明，二聚体界面部分地阻塞了Ub结合表面，尤其是在螺旋3的C末端，从而使UBA二聚化和Ub结合互斥。不寻常的是，单体UBA似乎是生物活性形式，而二聚体似乎是非活性形式。环1中的工程化点突变（E409K和G410K）显示出会破坏二聚体界面的稳定性，导致更高比例的结合单体，并且在NF-κB荧光素酶报告基因分析中，与wt-相比，NF-κB活性降低62。

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