Molecular basis of late onset ornithine transcarbamylase deficiency in male

我们报告了一群男性鸟氨酸转氨甲酰酶缺乏症（OTC）患者，发病年龄从青春期晚期到老年前期。在本研究中，我们发现了3例R4 OH突变患者。其中1例患者的肝脏标本中检测到R4 OH突变，而皮肤和成纤维细胞标本中的RFLP结果基本正常。提示该患者的R4 OH突变可能为体细胞突变。R4 OH突变在不同的民族中都有发现。这种突变可能是由于复发性突变，因为突变涉及CpG island.We已经表明，翻译后机制有关晚发型omithine转氨甲酰酶缺乏症患者轴承R4 OH或Y55 D突变。在本研究中，使用Cos 1细胞的表达分析表明，仅用该质粒转染的细胞和含有野生型cDNA的质粒转染的细胞的OTC活性分别为23.5（nmol/min/ml）和1955*140（n=3）。另一方面，R4 OH突变体cDNA和Y55 D cDNA的OTC活性分别为670*130（n=3）和627*124（n=3）。以β-半乳糖苷酶活性标准化的R4 OH突变体OTC和Y55 D突变体OTC的活性分别为正常OTC活性的28%和26%。当细胞裂解物经历5次冻融循环时，野生型OTC和Y55 D突变体OTC的活性没有变化，而R4 OH突变体OTC的活性下降到处理前野生型的6%。这些结果表明，R4 OH突变体OTC在物理上不稳定，并且比野生型酶降解更快。因此，一旦施加将促进突变酶的失活和降解的代谢负荷，R4 OH OTC活性可能被抑制至不相容的水平。

We had reported a cluster of male ornithine transcarbamylase deficiency (OTC) patients with onset from late adolescence to the presenile period. In this study, we discovered three patients who have R4OH mutation. We detected R4OH mutation in liver specimens from one of the patients with RFLP, but in fibroblast and skin specimens the RFLP showed almost normal. This result indicated that the R4OH mutation in the patient may be somatic mutation. The R4OH mutation has been discovered in different ethnic groups. This mutation may arise as a result of a recurrent mutation because the mutation involves a CpG island.We have showed that posttranslational mechanisms related late onset omithine transcarbamylase deficiency patients bearing R4OH or Y55D mutations. In this study, expression analysis using Cos 1 cells indicated that the OTC activities of cells transfected with the plasmid only, and the plasmid containing wild type cDNA were 23.5 (nmol/min/ml) and 1955*140 (n=3) respectively. On the other hand OTC activities with R4OH mutant cDNA, and Y55D cDNA were 670*130 (n=3) and 627*124 (n=3) respectively. The activities of R4OH mutant OTC and Y55D mutant OTC, as normalized for beta-galactosidase activity were 28% and 26% of the normal OTC activity respectively. When the cell lysates were subjected to five cycles of freezing and thawing, the activities of the wild type OTC and Y55D mutant OTC did not change, whereas the activity of the R4OH mutant OTC decreased to 6% of wild type that before treatment. These results indicated that the R4OH mutant OTC was physically unstable and was degraded more rapidly than the wild type enzyme. Thus, the R4OH OTC activity may be depressed to an incompatible level once a metabolic burden that would facilitate inactivation and degradation of the mutant enzyme is applied.