Kobayashi T, Ogawa H, Kasahara M, Shiozawa Z, Matsuzawa T
Department of Biochemistry, School of Medicine, Fujita Health University, Aichi, Japan.
Am J Hum Genet. 1995 Aug;57(2):284-91.
We describe here evidence of congenital enzyme mistargeting induced not by abnormalities in the signal sequence. We examined the molecular mechanism of hereditary ornithine aminotransferase (OAT) deficiency causing gyrate atrophy of the choroid and retina (GACR). Nucleotide sequencing of OAT cDNA generated from a GACR patient's mRNA revealed a single base change from C to G at position 268, resulting in an amino acid substitution of neutral Gln(CAA) with negatively charged Glu(GAA) at position 90 (Q90E). Immunohistochemical and transient expression analyses suggested expression of a defective labile OAT in the patient's tissues. However, high-level expression and immunocytochemical analyses elucidated that Q90E OAT (the patient's OAT) was localized within the limits of cytoplasmic free ribosomes in precursor form without any mitochondrial entry, indicating that the patient's precursor OAT was synthesized and rapidly degraded because of accumulation in the cytosol. It is interesting that, although the mutation site (Q90E) in this GACR patient's OAT was within the coding sequence of the mature protein, the precursor exhibited loss of mitochondrial targeting function. These findings suggest that not only the signal sequence but a critical part of the mature sequence plays an essential role in mitochondrial entry of the OAT precursor protein.
我们在此描述了并非由信号序列异常所诱导的先天性酶错误靶向的证据。我们研究了导致脉络膜和视网膜回旋性萎缩（GACR）的遗传性鸟氨酸氨基转移酶（OAT）缺乏症的分子机制。从一名GACR患者的mRNA生成的OAT cDNA的核苷酸测序显示，在第268位有一个从C到G的单碱基变化，导致第90位的中性谷氨酰胺（CAA）被带负电荷的谷氨酸（GAA）取代（Q90E）。免疫组织化学和瞬时表达分析表明，患者组织中存在有缺陷的不稳定OAT表达。然而，高水平表达和免疫细胞化学分析表明，Q90E OAT（患者的OAT）以前体形式定位于细胞质游离核糖体范围内，没有任何进入线粒体的情况，这表明患者的前体OAT由于在细胞质中积累而被合成并迅速降解。有趣的是，尽管该GACR患者的OAT中的突变位点（Q90E）位于成熟蛋白的编码序列内，但前体却表现出线粒体靶向功能丧失。这些发现表明，不仅信号序列，而且成熟序列的一个关键部分在OAT前体蛋白进入线粒体过程中起着至关重要的作用。
