TRANSIENT GLUCOSYLATION OF GLYCOPROTEINS

糖蛋白的瞬时糖基化

• 批准号: 6386001
• 负责人: ARMANDO JOSE PARODI
• 金额: $ 7.5万
• 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 2003-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6386001
• 关键词: Schizosaccharomyces pombe; Trypanosoma cruzi; calnexin; calreticulin; conformation; cysteine endopeptidases; endoplasmic reticulum; enzyme activity; enzyme mechanism; gene mutation; glycoprotein biosynthesis; glycoproteins; glycosylation; hexosyltransferase; laboratory rabbit; lectin; molecular chaperones; oligosaccharides; protein folding; protein structure function; protozoal antigen; uridine diphosphate glucose; virulence

Protein glycosylation is initiated in the endoplasmic reticulum (ER) by transfer of an oligosaccharide (Glc3Man9GlcNAc2) to nascent polypeptide chains. Processing of the oligosaccharide by glucosidases I and II yields unglucosylated oligosaccharides that are reglucosylated by the UDP-Glc:glycoprotein glucosyltransferase (GT) only if the protein moieties of glycoproteins are not properly folded, as GT behaves as a sensor of glycoprotein conformations. Interaction of monoglucosylated glycoproteins (created by partialdeglucosylation of the transferred compound or by GT-mediated reglucosylation) with ER lectins (calnexin and calreticulin) facilitates acquisition of the proper tertiary structures and prevents secretion of not yet properly folded species. Folding facilitation mediated by the interaction of monoglucosylated glycoproteins and ER lectins is necessary for production of infective viral particles (vesicular stomatitis virus, HIV, hepatitis B virus) and also for the viability of yeasts when under severe ER stress conditions. The long term objective of the proposal is to gain thorough information of the structural features leading to the monoglucosylated glycoprotein-lectin interaction as this knowledge will undoubtedly provide useful information for understanding and/or preventing production of the etiological agents of several diseases. Within this long term objective, the Specific Aims of the proposed research are: a) to define the structural elements exclusively exposed in misfolded glycoproteins whose recognition is required for GT-mediated glucosylation; b) to define GT domains responsible for substrate donor (UDP-Glc), and substrate acceptor (N-oligosaccharide) recognition and for the exclusive glucosylation of misfolded species and c) to study the possibility that folding facilitation mediated by the interaction of cruzipain, a cysteine proteinase from the protozoan parasite Trypanosoma cruzi and ER lectins could be absolutely necessary for parasite infectivity. T. cruzi is the causative agent of a disease endemic in Latin America (Chagas's disease) that affects about 16 million people. Cruzipain has been identified as one of the T. cruzi virulence factors and has been found to be the only glycoprotein interacting with ER lectins in the protozoon.

通过将寡糖 (Glc3Man9GlcNAc2) 转移至新生多肽链，在内质网 (ER) 中启动蛋白质糖基化。 通过葡糖苷酶 I 和 II 加工寡糖会产生未葡糖基化的寡糖，仅当糖蛋白的蛋白质部分未正确折叠时，这些寡糖才会被 UDP-Glc:糖蛋白葡糖基转移酶 (GT) 重新葡糖基化，因为 GT 充当糖蛋白构象的传感器。 单糖基化糖蛋白（通过转移化合物的部分脱糖基化或通过 GT 介导的重新糖基化产生）与 ER 凝集素（钙联蛋白和钙网蛋白）的相互作用有助于获得正确的三级结构并防止尚未正确折叠的物质的分泌。 由单糖基化糖蛋白和内质网凝集素相互作用介导的折叠促进对于感染性病毒颗粒（水泡性口炎病毒、艾滋病毒、乙型肝炎病毒）的产生以及酵母在严重内质网应激条件下的生存能力是必需的。 该提案的长期目标是获得导致单糖基化糖蛋白-凝集素相互作用的结构特征的全面信息，因为这些知识无疑将为理解和/或预防几种疾病的病原体的产生提供有用的信息。 在这一长期目标中，拟议研究的具体目标是： a) 定义仅在错误折叠糖蛋白中暴露的结构元件，这些元件的识别是 GT 介导的糖基化所必需的； b) 定义负责底物供体 (UDP-Glc) 和底物受体 (N-寡糖) 识别以及错误折叠物种的专有糖基化的 GT 结构域，以及 c) 研究由 cruzipain 相互作用介导的折叠促进的可能性，来自原生动物寄生虫克氏锥虫的半胱氨酸蛋白酶和 ER 凝集素对于寄生虫的感染性可能是绝对必要的。 克氏锥虫是拉丁美洲流行病（恰加斯病）的病原体，影响约 1600 万人。 Cruzipain 已被鉴定为 T. cruzi 毒力因子之一，并且被发现是与原生动物中 ER 凝集素相互作用的唯一糖蛋白。