Identifying understudied protein-related glycoproteome disruption in Congenital Disorders of Glycosylation

项目总结/摘要 先天性糖基化障碍（CDG）是一组罕见的神经代谢遗传性疾病， 破坏糖基化，即蛋白质中添加糖结构。许多CDG基因编码研究不足的， 潜在的药物蛋白质。糖基化作为最丰富的翻译后修饰（PTM）， 产生巨大的生物变异性并介导基本的生物过程。CDG患者表现出 多器官功能障碍，这往往是严重的早期死亡率。CDG的病理生理学归因于 破坏蛋白质糖基化;然而，负责蛋白质糖基化的低糖基化蛋白质的特定身份 大多数疾病的表现是未知的。与理解病理机制相关的关键未满足需求 CDG患者仍有有效的治疗方法。在这里，我们建议利用基于质谱的 技术，以评估具有致病性变异的CDG患者样本中的糖蛋白质组学破坏， 可药用的、未充分研究的蛋白质（SLC 35 A2、SLC 35 A3和SLC 39 A8）。了解遗传缺陷如何在 编码这些蛋白质的基因破坏了糖基化，这将为我们提供新的科学见解。 CDG疾病表现的病理生理学，可能建议新的治疗策略，并将告知 糖基化的正常功能和这些研究不足的蛋白质。 这些选定的蛋白质的可药用性主要是由于它们作为底物转运蛋白的作用， 这些蛋白质使聚糖亚基能够被摄取并定位于内质网和高尔基体 用于聚糖合成和成熟的装置。大多数致病性CDG变体是亚型突变， 导致转运蛋白底物的可用性增加可以克服蛋白质功能障碍的假设， 恢复糖基化并减轻疾病表现。除了评估糖蛋白组破坏外 在来自受影响个体的样本中，本提案将测试转运蛋白底物补充剂是否 可以使疾病状态的糖蛋白质组破坏正常化并克服潜在的蛋白质功能障碍。 本研究将使用最先进的糖蛋白组学技术和临床相关患者样本 阐明未充分研究的CDG转运蛋白的功能。仪器仪表的先驱性进步， 实验方法和计算方法在糖蛋白质组学的鉴定， 糖蛋白质组破坏终于实现。这些研究将鉴定低糖基化蛋白， 并生成数据以解决底物补充是否 在这些CDG可能解决疾病表现的根本原因。的知识和数据 这些研究所产生的结果将是申请资金和开展未来研究的关键， 治疗由SLC 35 A2、SLC 35 A3和SLC 39 A8中的缺陷引起的CDG病症。

PROJECT SUMMARY/ABSTRACT The congenital disorders of glycosylation (CDG) are a group of rare neurometabolic genetic diseases that disrupt glycosylation, the addition of sugar structures to proteins. Many CDG genes encode understudied and potentially druggable proteins. As the most abundant post-translational modification (PTM), glycosylation generates immense biological variability and mediates fundamental biological processes. CDG patients exhibit multiorgan dysfunction, which is often severe with early mortality. Pathophysiology of CDG is attributed to disrupted protein glycosylation; however, the specific identities of hypoglycosylated proteins responsible for most disease manifestations are unknown. Critical unmet needs related to understanding pathomechanisms and effective therapies remain for CDG patients. Here, we propose to utilize mass spectrometry-based technologies to evaluate glycoproteomic disruptions in CDG patient samples with pathogenic variants in druggable, understudied proteins (SLC35A2, SLC35A3, and SLC39A8). Understanding how genetic defects in genes encoding these proteins disrupts glycosylation will provide new scientific insights into the pathophysiology of CDG disease manifestations, may suggest novel treatment strategies, and will inform normal function of glycosylation and these understudied proteins. The druggability of these selected proteins is largely due to their roles as substrate transporters, through which the proteins enable uptake and localization of glycan subunits to the endoplasmic reticulum and Golgi apparatus for glycan synthesis and maturation. Most pathogenic CDG variants are hypomorphic mutations, leading to the hypothesis that increased availability of transporter substrate may overcome protein dysfunction, restore glycosylation, and mitigate disease manifestations. In addition to evaluating glycoproteomic disruption in specimens from affected individuals, this proposal will test whether transporter substrate supplementation can normalize the glycoproteomic disruption of the disease state and overcome underlying protein dysfunction. This study will use using state-of-the-art glycoproteomics technologies and clinically relevant patient samples to elucidate functions of understudied CDG transporter proteins. Pioneering advances in instrumentation, experimental methodologies, and computational approaches in glycoproteomics make identification of glycoproteome disruption finally achievable. These studies will identify hypoglycosylated proteins and glycosites in patient tissues and generate data to address the question of whether substrate supplementation in these CDG may address the underlying causes of disease manifestations. The knowledge and data generated from these studies will be pivotal for applying for funding and carrying out future studies aimed at treating CDG disorders caused by defects in SLC35A2, SLC35A3, and SLC39A8.