The relationship between the intracellular stability of tyrosine hydroxylase and the neurodegeneration on Parkison's disease

α -突触核蛋白突变引起的泛素-蛋白酶体系统异常与帕金森病的发病机制有关。最近，酪氨酸羟化酶（TH）被发现是在体外重组系统中与泛素结合的底物，并且在网状细胞裂解系统中被蛋白酶体部分降解。TH的n端被认为位于分子表面，并被认为具有易受蛋白酶（如钙蛋白酶）影响的柔性构象。因此，TH的n端被认为是细胞内蛋白水解的靶区，如发生在泛素-蛋白酶体系统中。总之，这些结果表明TH的n端可能在TH分子的稳定性以及细胞中TH催化活性的调节中发挥关键作用。本研究设计了一个实验方案来证明n端对TH分子稳定性的影响。首先，我们澄清了n端区域的缺失增强了人类TH型1分子的细胞内稳定性，并且在产生n端缺失突变体的细胞中多巴胺的积累可归因于TH分子的增强稳定性。此外，计算机辅助分析人类TH 1型的空间结构，鉴定出5个新识别的PEST基序，它们是蛋白酶体的靶序列，其中一个位于Met^1- lys ^<12>的n端序列。我们相信本研究为了解哺乳动物细胞中TH蛋白的稳定性和/或帕金森病的发病机制提供了一些信息。

The abnormality of ubiquitin-proteasome system caused by the mutations of alpha-synuclein has been implicated in the pathogenesis of Parkinson's disease. Recently, tyrosine hydroxylase (TH) was found to be a substrate for conjugation to ubiquitin in a reconstituted in vitro system and to be partially degraded by proteasomes in a reticulocyte lysate system. The N-terminus of TH is supposed to be located on the surface of the molecule, and it is presumed to possess flexible conformation susceptible to proteases such as calpain. Therefore, the N-terminus of TH is thought to be a target region for intracellular proteolysis such as occurs in the ubiquitin-proteasome system. Collectively, these results suggest that the N-terminus of TH might play a critical role in the stability of the TH molecule as well as in the regulation of TH catalytic activity in cells.In this research, an experimental scheme was designed in order to prove the influence of the N-terminus on the stability of the TH molecule. At first, we clarified that the deletion of the N-terminus region augmented the intracellular stability of the human TH type 1 molecule and that the accumulation of dopamine in the cells producing the N-terminus-deleted mutants can be attributed to augmented stabilization of the TH molecule. Moreover, computer-assisted analysis of the spatial configuration of human TH type 1 identified 5 newly recognized PEST motifs, the target sequence for proteasome, and one of which was located in the N-terminus sequence of Met^1-Lys^<12>. We believe that this study provides some information to understand the stability of TH protein in mammalian cells and/or the pathogenesis of Parkinson's disease.