Posttranslational Modifications of Proteins during Intracellular Transport

细胞内运输过程中蛋白质的翻译后修饰

基本信息

批准号：
02670117
负责人：
IKEHARA Yukio
金额：
$ 1.54万
依托单位：
Fukuoka University
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1990
资助国家：
日本
起止时间：
1990 至 1991
项目状态：
已结题

项目摘要

1. -Identification of furin involved in the processing of proproteins1) Primary structure of furin deduced from the cDNA : The cDNA of furin was isolated from human and rat liver cDNA libraries. The cDNA sequences indicate that the human and rat furins consist of 794 and 793 amino acids, respectively. The primary structure of each furin shows high homology to that of the yeast Kex2 protease, a prohormone-processing enzyme, and contains three potential sites for glycosylation and a transmenbrane domain at a COOH-terminal region.2) Intracellular localization and processing activity of furin : The furin cDNA was transfected into COS-1 cells. Immunoelectron-microscopical observations revealed that furin is localized in the Golgi complex. When albumin or complement C3 was expressed by transfection of each cDNA, each proform was completely processed into a mature form by furin co-expressed in the same cells. These results indicate that furin, localized in the Golgi complex, is an endoproteas … More e involved in the processing of proforms of secretory proteins.2. Biosynthesis, processing and transport of membrane proteins1) GPI-anchored proteins : We have established that the ectoenzyme alkaline phosphatase (ALP) is initially synthesized as a proform having a hydrophobic sequence at the COOH terminus, which is immediately cleaved and replaced by a glycosyl-phosphatidylinositol (GPI) anchor, and the GPI-anchored form is transported to the cell surface. When the biosynthesis of GPI was completely blocked by 2-fluoro-2-deoxyglucose, the newly synthesized ALP remained in the endoplasmic reticulum as the proform, which was then gradually converted to a soluble form by proteolytic removal of the COOH-terminal segment, followed by its secretion. The results indicate that the GPI anchoring is prerequisite for ALP to be transported to the cell surface.2) An enzyme defect due to proteolytic degradation in the endoplasmic reticulum (ER) : The ectoenzyme dipeptidyl peptidase IV (DPPIV) was found to be defective in a rat substrain. Cloning and sequencing of the DPPIV cDNA identified a point mutation which leads to substitution of Gly^<633>->Arg at the active site sequence of DPPIV. In addition, the mutant DPPIV was found to undergo rapid degradation in the ER without being transported to the cell surface. Thus, it is concluded that the rapid degradation of the mutant DPPIV in the ER results, in the enzyme defect in the affected rat. Less

1。-识别弗林参与先知加工的识别1）从cDNA推导出的脂蛋白的主要结构：从人和大鼠肝cDNA文库中分离出脂蛋白的cDNA。 cDNA序列表明人和大鼠芦丝分别由794和793个氨基酸组成。每种呋喃蛋白的主要结构与酵母Kex2蛋白酶的同源性很高，一种激素加工酶，并包含三个潜在的糖基化位点，并且在COOH-末端区域都包含糖基化的三个潜在部位。免疫电子显微镜观察结果表明，Furin位于高尔基体复合物中。当专辑或完成C3通过转染每个cDNA表达时，每种利润通过在同一细胞中共表达的Furin完全处理成成熟的形式。这些结果表明，在高尔基体配合物中局部的弗林是一个内前肢……更多地参与了秘密蛋白质的加工。2。 Biosynthesis, processing and transport of membrane proteins1) GPI-anchored proteins: We have established that the ectoenzyme alcoholine phosphatase (ALP) is initially synthesized as a hydrophobic sequence at the COOH terminus, which is immediately cleaved and replaced by a Glycosyl-phosphatidylinositol (GPI) anchor, and the GPI-anchored form is运输到细胞表面。当2-氟-2-脱氧葡萄糖完全阻断GPI的生物合成时，新合成的ALP作为型构型保留在内质网中，然后逐渐通过对COOH-末端段的蛋白水解去除，然后将其分泌。结果表明，GPI锚定是将ALP转运到细胞表面的先决条件。2）由于内胞质网状（ER）中蛋白水解降解引起的酶缺损：在大鼠底因中发现了二肽二肽基肽IV（DPPIV）的均二酰基二肽基IV（DPPIV（DPPIV）。 DPPIV cDNA的克隆和测序鉴定了一个点突变，该突变导致在DPPIV的活性位点序列上取代Gly^<633> - > arg。此外，发现突变型dppiv在ER中经历了快速降解，而无需转运到细胞表面。这就是结论，在受影响大鼠的酶缺陷中，ER结果中突变体DPPIV的快速降解。较少的