Protein instability and functional defects caused by mutations of dihydro-orotate dehydrogenase in Miller syndrome patients.
Protein instability and functional defects caused by mutations of dihydro-orotate dehydrogenase in Miller syndrome patients.
复制标题
蛋白质的不稳定性和功能缺陷是由米勒综合征患者中二氢 - 二氢甲酸脱氢酶突变引起的。
DOI：
10.1042/bsr20120046
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发表时间：
2012-12
影响因子：
4
通讯作者：
Kang D
中科院分区：
文献类型：
--
作者：
Fang J;Uchiumi T;Yagi M;Matsumoto S;Amamoto R;Saito T;Takazaki S;Kanki T;Yamaza H;Nonaka K;Kang D
米勒综合征是一种隐性遗传性疾病，其特征是轴后肢端面骨发育不全。它是由DHODH（二氢乳清酸脱氢酶）基因功能障碍引起的，DHODH基因编码嘧啶从头生物合成途径中的关键酶，并定位于线粒体膜间隙。我们研究了DHODH的三个错义突变G202 A、R346 W和R135 C的后果，这些突变先前在米勒综合征患者中被发现。首先，我们建立了稳定表达具有米勒综合征致病突变：G202 A、R346 W和R135 C的DHODH的HeLa细胞系。这三种突变蛋白保留了适当的线粒体定位的基础上，免疫组化和线粒体亚分级研究。G202A、R346W DHODH蛋白质显示降低的蛋白质稳定性。另一方面，第三个R135C，其中的突变位于泛醌结合位点，是稳定的，但没有酶活性。总之，G202 A和R346 W突变导致蛋白质稳定性缺陷，R135 C突变不影响稳定性，但损害底物诱导的酶活性，表明DHODH活性的损害与米勒综合征表型相关。
Miller syndrome is a recessive inherited disorder characterized by postaxial acrofacial dysostosis. It is caused by dysfunction of the DHODH (dihydroorotate dehydrogenase) gene, which encodes a key enzyme in the pyrimidine de novo biosynthesis pathway and is localized at mitochondria intermembrane space. We investigated the consequence of three missense mutations, G202A, R346W and R135C of DHODH, which were previously identified in patients with Miller syndrome. First, we established HeLa cell lines stably expressing DHODH with Miller syndrome-causative mutations: G202A, R346W and R135C. These three mutant proteins retained the proper mitochondrial localization based on immunohistochemistry and mitochondrial subfractionation studies. The G202A, R346W DHODH proteins showed reduced protein stability. On the other hand, the third one R135C, in which the mutation lies at the ubiquinone-binding site, was stable but possessed no enzymatic activity. In conclusion, the G202A and R346W mutation causes deficient protein stability, and the R135C mutation does not affect stability but impairs the substrate-induced enzymatic activity, suggesting that impairment of DHODH activity is linked to the Miller syndrome phenotype.