Purpose

Autosomal dominant acute necrotizing encephalopathy (ADANE) is caused by missense mutations in the gene encoding Ran-binding protein 2 (RANBP2), a nuclear pore protein regulating mitochondrial localization and function. Previous studies have found that RANBP2 binds to COX11 and suppresses its inhibitory activity over hexokinase1. To further elucidate mitochondrial dysfunction in ADANE, we analyzed the interaction between mutated RANBP2 and COX11.

Methods

We extracted cDNA from a patient and constructed pGEX wild-type or mutant-type vectors including RANBP2 c.1754C>T, the commonest variant in ADANE. We transformed E. coli competent cells with the vectors and had them express GST-RANBP2 recombinant protein, and conducted a pull-down assay of RANBP2 and COX11.

Results

The amount of COX11 bound to mutated RANBP2 was significantly smaller than that bound to the wild-type RANBP2.

Conclusion

Mutated RANBP2 had an attenuated binding ability to COX11. Whether this change indeed decreases ATP production remains to be further explored.

中文翻译：

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导致常染色体显性遗传的急性坏死性脑病的 RANBP2 突变减弱了其与 COX11 的相互作用

目的

常染色体显性遗传急性坏死性脑病 (ADANE) 是由编码 Ran 结合蛋白 2 (RANBP2) 的基因错义突变引起的，RANBP2 是一种调节线粒体定位和功能的核孔蛋白。先前的研究发现，RANBP2 与 COX11 结合并抑制其对己糖激酶1 的抑制活性。为了进一步阐明 ADANE 中的线粒体功能障碍，我们分析了突变的 RANBP2 和 COX11 之间的相互作用。

方法

我们从患者身上提取了 cDNA，并构建了 pGEX 野生型或突变型载体，包括RANBP2 c.1754C>T，ADANE 中最常见的变体。我们用载体转化大肠杆菌感受态细胞并使其表达GST-RANBP2重组蛋白，并对RANBP2和COX11进行下拉测定。

结果

与突变的 RANBP2 结合的 COX11 的量明显小于与野生型 RANBP2 结合的量。

结论

突变的 RANBP2 与 COX11 的结合能力减弱。这种变化是否确实降低了 ATP 的产生仍有待进一步探索。