Transient Glucosylation of Glycoproteins

糖蛋白的瞬时糖基化

• 批准号: 6893421
• 负责人: ARMANDO JOSE PARODI
• 金额: $ 8.1万
• 依托单位: FUNDACION INSTITUTO LELOIR
• 依托单位国家: 美国
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-08-15 至 2008-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6893421
• 关键词: Golgi apparatus; Schizosaccharomyces pombe; calnexin; calreticulin; conformation; cysteine endopeptidases; endoplasmic reticulum; enzyme activity; enzyme mechanism; glycoprotein biosynthesis; glycoproteins; glycosylation; hexosyltransferase; intracellular transport; laboratory mouse; laboratory rabbit; laboratory rat; molecular chaperones; oligosaccharides; protein folding; protein structure function; protein transport; uridine diphosphate glucose

DESCRIPTION (provided by applicant): The quality control of glycoprotein folding in the endoplasmic reticulum (ER) involves the interplay of a glucosyltransferase (GT) that only glycosylates not properly folded protein conformers, glucosidase lI (GII) that removes residues added by GT and two ER resident lectins (calnexin, CNX and calreticulin, CRT) that specifically recognize monoglucosylated glycoproteins. This mechanism prevents exit of not properly folded glycoproteins to the Golgi and enhances glycoprotein folding efficiency. We have recently obtained evidence indicating that GT recognizes hydrophobic amino acid patches exposed in molten globule-like conformations displayed at the last glycoprotein folding stages. The purpose of Specific Aim I is to study, at the molecular level, glycoprotein interaction with GT-CNX/CRT and other ER chaperones, from the time the polypeptide moiety emerges into the ER lumen until an oligomer complex is formed. As UDP-Glc is GT substrate donor, the glucosylating reaction produces UDP as one of its reaction products. Nucleoside diphosphates generated by glycosyltransferases in the secretory pathway must be converted into monophosphates to relieve inhibition of the transferring enzymes and to provide substrates for antiport transport systems by which entrance of nucleotide sugars from the cytosol into the secretory pathway lumen is coupled to exit of nucleoside monophosphate. We have recently obtained evidence suggesting that in the yeast Schizosaccharomyces pombe GT-generated UDP might be hydrolyzed by nucleoside diphosphatases occurring not in the ER but in the Golgi apparatus. Specific Aim II deals with the possibility that vesicular transport between the ER and cis Golgi cisternae might carry not only the macromolecular components of the quality control mechanism (GT, GII, CNX and CRT have ER retrieval sequences at their C-termini) but also UDP in the anterograd movement to be hydrolyzed by Golgi GDPases/UDPases and LIMP in the retrograd movement. Finally, Specific Aim III deals on the mechanism, intimately intertwined with that of quality control, by which cells distinguish between folding intermediates that are in the process of productive folding and irremediably misfolded glycoproteins that have to be diverted to proteasomal degradation.

描述（由申请人提供）：内质网（ER）中糖蛋白折叠的质量控制涉及仅糖基化未正确折叠的蛋白质构象异构体的葡萄糖基转移酶（GT）、去除由GT添加的残基的葡萄糖苷酶II（GII）和两种内质网驻留凝集素（钙联蛋白，CNX 和钙网蛋白，CRT）专门识别单葡萄糖基化糖蛋白。这种机制可以防止未正确折叠的糖蛋白退出高尔基体并提高糖蛋白折叠效率。我们最近获得的证据表明，GT 可以识别在糖蛋白最后折叠阶段显示的熔球状构象中暴露的疏水性氨基酸斑块。具体目标 I 的目的是在分子水平上研究糖蛋白与 GT-CNX/CRT 和其他 ER 分子伴侣的相互作用，从多肽部分进入 ER 腔直至形成寡聚物复合物。由于UDP-Glc是GT底物供体，因此糖基化反应产生UDP作为其反应产物之一。分泌途径中糖基转移酶产生的核苷二磷酸必须转化为单磷酸，以减轻对转移酶的抑制，并为逆向转运系统提供底物，通过该系统，核苷酸糖从胞质溶胶进入分泌途径腔与核苷单磷酸的退出相偶联。我们最近获得的证据表明，在酵母裂殖酵母 GT 中产生的 UDP 可能会被核苷二磷酸酶水解，核苷二磷酸酶不存在于内质网中，而是存在于高尔基体中。具体目标 II 涉及 ER 和顺式高尔基体池之间的囊泡运输可能不仅携带质量控制机制的大分子成分（GT、GII、CNX 和 CRT 在其 C 端具有 ER 检索序列），还携带 UDP在顺行运动中被高尔基体 GDPases/UDPases 和 LIMP 在逆行运动中水解。最后，特定目标 III 涉及与质量控制密切相关的机制，细胞通过该机制区分处于生产性折叠过程中的折叠中间体和必须转向蛋白酶体降解的不可挽回的错误折叠糖蛋白。