Molecular mechanism of substrate recognition of the ubiquitin-ligases that recognize sugar chain

识别糖链的泛素连接酶底物识别的分子机制

基本信息

批准号：
15580085
负责人：
YOSHIDA Yukiko
金额：
$ 2.37万
依托单位：
TOKYO MRTROPOLITAN ORGANIZATION FOR MEDICAL RESEARCH
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2003
资助国家：
日本
起止时间：
2003 至 2004
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-15580085/
关键词：
N-glycan Ubiquitin-ligase Lectin Endoplasmic reticulum associated degaradation (ERAD)

项目摘要

In an attempt to find novel biological functions for N-glycans, we screened mouse brain extracts for proteins bound to various sugar probes, and found Fbx2 an F-box protein, binds specifically to proteins attached with N-linked high-mannose type oligosaccharides, and subsequently contributes to ubiquitylation of N-glycosylated proteins. We propose that SCF^<Fbx2> ubiquitylaltes N-glycosylated proteins, which are translocated from the ER to the cytosol by the quality control mechanism. In this study, we found another F-box protein, Fbs6b that recognize N-glycans. Both Fbx2 and Fbx6b interacted with glycoproteins containing high-mannose oligosaccharides, whose protein modification occurs in the ER. X-ray crystallographic and nuclear magnetic resonance (NMR) studies of the substrates-binding domain of Fbx2 revealed that Fbx2 recognized the inner chitobiose of high-mannose oligosaccharides by a small hydrophobic pocket located at the top of the β-barrel. Both Fbx2 and Fbx6b proteins interacted with denatured glycoproteins, which were modified with not only high-mannose but also complex-type oligosaccharides, more efficiently than native proteins. Given that Fbs proteins interact with innermost chitobiose in N-glycans, we propose that Fbs proteins distinguish native from unfolded glycoproteins by sensing the exposed chitobiose structure.

为了寻找 N-聚糖的新生物学功能，我们筛选了小鼠大脑提取物中与各种糖探针结合的蛋白质，并发现 Fbx2 是一种 F-box 蛋白，可与 N-连接的高甘露糖型寡糖所附着的蛋白质特异性结合，并随后促进 N-糖基化蛋白的泛素化，我们提出 SCF^<Fbx2> 泛素化 N-糖基化蛋白，这些蛋白是从 N-糖基化蛋白易位的。通过质量控制机制将内质网转移到细胞质中。在这项研究中，我们发现了另一种识别 N-聚糖的 F-box 蛋白 Fbs6b，Fbx2 和 Fbx6b 都与含有高甘露糖寡糖的糖蛋白相互作用，其蛋白修饰发生在内质网中。 Fbx2 底物结合域的 X 射线晶体学和核磁共振 (NMR) 研究表明，Fbx2 识别内部壳二糖Fbx2 和 Fbx6b 蛋白通过位于 β-桶顶部的小疏水口袋与高甘露糖寡糖相互作用，这些蛋白不仅用高甘露糖修饰，而且还用复合型寡糖修饰，比天然蛋白更有效。鉴于 Fbs 蛋白与 N-聚糖中最内部的壳二糖相互作用，我们建议 Fbs 蛋白通过感测暴露的糖蛋白来区分天然糖蛋白和未折叠糖蛋白。壳二糖结构。