PEPTIDE:N-GLYCANASE

肽：N-聚糖酶

• 批准号: 7726232
• 负责人: HERMANN SCHINDELIN
• 金额: $ 0.55万
• 依托单位: BROOKHAVEN SCIENCE ASSOC-BROOKHAVEN LAB
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-09-18 至 2009-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7726232
• 关键词: ATP phosphohydrolase; Adopted; Binding; Biochemical; C-terminal; Catalytic Domain; Cell Survival; Complex; Computer Retrieval of Information on Scientific Projects Database; Data; Data Collection; Degradation Pathway; Endoplasmic Reticulum; Enzymes; Eukaryota; Eukaryotic Cell; Funding; Grant; Institution; Length; Modeling; Mus; N-terminal; Peptide N-Glycosidase; Polysaccharides; Process; Protein Family; Proteins; Reporting; Research; Research Personnel; Resources; Solutions; Source; Structure; Time; United States National Institutes of Health; interest; multicatalytic endopeptidase complex; novel; protein degradation; protein misfolding; sugar; yeast protein

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The endoplasmic reticulum-related protein degradation (ERAD) pathway, where misfolded proteins in the ER are transported through the retro-translocon, ubiquinated, and degraded by the proteasome, is essential for cell viability. Before proteasome degradation, the N-glycan on the ERAD substrates is processed by the Peptide:N-glycanase (PNGase). We tried to illustrate the function of PNGase by studying its structure. We recently solved the structure of the catalytic domain of mouse PNGase (mPNGase), which shows a similar fold as the yeast protein. In addition to the catalytic domain, PNGase in higher eukaryotes has both N- and C-terminal extensions. Our structural and biochemical studies on the C-terminal domain of mPNGase identified it as a sugar-recognizing motif. However, the function of the N-terminal domain is unknown. The N-terminal domain has no sequence similarity with other proteins and may adopt a novel fold. Recently it was reported that mPNGase directly interacts with P97/VCP, an AAA ATPase family protein. We proposed an escort model in which P97 functions as a protein interacting platform to present misfolded proteins from the ER to mPNGase and ubiquitinating enzymes. Our biochemical data showed that the N-terminal domain of mPNGase binds to the C-terminal region of P97. This is the first time that a protein-interacting function of the mPNGase N-terminal domain has been established. This is very interesting since it was generally believed that P97 interacts with its effector proteins through its N-terminal domain. The structure of full length P97 has been solved, however, the C-terminal region could not be visualized in the structure. To investigate the function of the N-terminal domain of mPNGase and the details of its interaction with P97, crystallographic studies on these proteins and their complexes are being undertaken. We have recently obtained crystals of the N-terminal domain in its apo-state and in complex with a fragment of P97 and expect them to be ready for data collection and structure solution by the time the RapiData course starts.

该子项目是利用该技术的众多研究子项目之一 资源由 NIH/NCRR 资助的中心拨款提供。子项目及 研究者 (PI) 可能已从 NIH 的另一个来源获得主要资金， 因此可以在其他 CRISP 条目中表示。列出的机构是 对于中心来说，它不一定是研究者的机构。 内质网相关蛋白降解 (ERAD) 途径对于细胞活力至关重要，其中 ER 中错误折叠的蛋白质通过逆转录转位子转运、泛素化并被蛋白酶体降解。在蛋白酶体降解之前，ERAD 底物上的 N-聚糖由肽：N-聚糖酶 (PNGase) 处理。我们试图通过研究 PNGase 的结构来说明它的功能。我们最近解析了小鼠 PNGase (mPNGase) 催化结构域的结构，它显示出与酵母蛋白相似的折叠。除了催化结构域外，高等真核生物中的 PNGase 还具有 N 端和 C 端延伸。我们对 mPNGase C 端结构域的结构和生化研究将其鉴定为糖识别基序。然而，N 端结构域的功能尚不清楚。 N端结构域与其他蛋白质没有序列相似性，并且可能采用新的折叠。 最近有报道称 mPNGase 直接与 AAA ATPase 家族蛋白 P97/VCP 相互作用。我们提出了一个护送模型，其中 P97 作为蛋白质相互作用平台，将错误折叠的蛋白质从 ER 呈现给 mPNGase 和泛素化酶。我们的生化数据表明 mPNGase 的 N 端结构域与 P97 的 C 端区域结合。这是首次建立 mPNGase N 末端结构域的蛋白质相互作用功能。这非常有趣，因为人们普遍认为 P97 通过其 N 端结构域与其效应蛋白相互作用。全长P97的结构已经解析，然而，C端区域在结构中无法可视化。为了研究 mPNGase N 末端结构域的功能及其与 P97 相互作用的细节，正在对这些蛋白质及其复合物进行晶体学研究。我们最近获得了处于 apo 状态且与 P97 片段复合的 N 端结构域晶体，并期望在 RapiData 课程开始时它们已准备好用于数据收集和结构解决方案。