Studies on catalytic mechanism of prolyl endopeptidase by genetic method and biological significance of it in the brain

克隆了脑膜败血黄杆菌、嗜水气单胞菌和牛脑的脯氨酰内肽酶基因[EC 3.4.21.26]和大肠杆菌的蛋白酶II基因。在这四个酶之间观察到一级结构高度保守。令人惊讶的是，Prolyl内肽酶的C末端区域与下列蛋白质有显著的同源性：猪和大鼠的酰基氨基酸释放酶，人蛋白3p21，大鼠和人的二肽氨基肽酶IV，酵母二肽氨基肽酶B，以及乳酸乳球菌的X-Prolyl二肽氨基肽酶。催化三联体的氨基酸残基非常保守。这些酶在寡肽上也有共同的特征。因此，Prolyl寡肽家族的存在被认为是一个新的丝氨酸内肽家族。淀粉样蛋白b/A4肽是阿尔茨海默病患者老年斑和脑血管淀粉样蛋白中典型的组成部分之一。本研究首次用免疫组织化学染色的方法，对加速衰老小鼠(SAM)体内的脯氨酰内肽酶的分布进行了研究。我还发现，Pro内肽酶和淀粉样蛋白b/A4肽在海马区分布密切相关。这些结果提示，Pro内肽酶参与了SAM脑内淀粉样蛋白沉积的形成过程。

The prolyl endopeptidase [EC 3.4.21.26] genes of Flavobacterium meningosepticum, A.hydrophila, and bovine brain, and protease II gene from E.coli were cloned. A high conservation of primary structure is observed among these four enzumes. Surprisingly, C-terminal regions of prolyl endopeptidase show significant homology to following proteins : pig and rat acyl-amino acid releasing enzymes, human protein 3P21, rat and human dipeptidyl aminopeptidases IV, yeast dipeptidyl aminopeptidase B, and X-prolyl dipeptidyl aminopeptidase from Lactococcus lactis. The amino acid residues of the catalytic triad are well conserved. These enzymes have also common characteristics on oligopeptides. Thus, presence of prolyl oligpeptidase family was proposed as a new family of serine endopeptidases.Amyloid b/A4 peptide is one of the integral components that are typically manifested in senile plaques and cerebrovascular amyloids in the patients with Alzheimer's disease. I ascertained for the first time the distribution of prolyl endopeptidase in senescence accelerated mouse (SAM) by the immunostaining method. I also found that prolyl endopeptidase and amyloid b/A4 peptide distributed with close relationships in the hippocampus. These result suggest that prolyl endopeptidase is involved in the processes to form amyloid deposition in the brain of SAM.