Developing chemoenzymatic strategies, enzymes, and kits for accessible and affordable gangliosides

项目摘要 神经节苷脂，类似于细胞中的生物触角，参与多种生物的关键作用。 过程(即病原体感染、细胞间通讯、炎症)。神经保护和神经修复 神经节苷脂的功能已被探索用于帕金森病和其他疾病的治疗 神经退行性疾病。尽管神经节苷脂有如此重要的作用，但神经节苷脂的获取途径有限。 目前商业上可获得的神经节苷脂大多来自动物的大脑，这可能是一种 对未来治疗应用的关注。神经节苷脂的这种有限的可得性进一步受到 基于不同的糖和脂肪酸的多样性。这里介绍的技术涵盖了 经过近十年的研究，在单个反应容器中合成了复杂的神经节苷脂，然后 一种简单的纯化技术。这种生物合成工具打开了急需的神经节苷脂在 研究并支持其治疗用途。在这项快速通道提案中，我们将解决以下几个主要挑战 将这一原型一锅多酶(OPME)技术转化为大规模商业化。第一 挑战是制造原料(糖基转移酶、补充酶和 乳酸鞘氨醇)，以降低生产神经节苷脂的成本。对现有的其他改进 糖基转移酶将通过改进催化作用进一步降低生产神经节苷脂的成本负担 酶的效率。最后，将探讨OPME的可伸缩性，以制造各种 神经节苷脂。一项研究将导致几种商业产品的小型化，从纯化的酶到 糖基化试剂盒和几个具有很高商业价值的关键神经节苷脂。GM1神经节苷脂已被报道 为了延缓帕金森病的发病，临床试验可能需要不含动物的神经节苷脂。一台微型的 试剂盒将作为研究人员合成他们自己定制的神经节苷脂的工具。这个 糖基转移酶和OPME试剂盒允许广泛的科学界访问这些简单的合成 其他寡糖、糖偶联物和糖蛋白的工具。通过扩大这一OPME并降低 这些产品的成本，将获得关于神经节苷脂生物学功能的更多信息，这将 加快发展以神经节苷脂为基础的诊疗。一个正反馈循环，其中 更低的成本将增加采购量，这将满足更多的研究，并将导致进一步的增长 糖基转移酶、OPME试剂盒和高纯度、无动物神经节苷脂的市场。

Project Summary Gangliosides, analogous to biological antennae in cells, are involved in critical roles across multiple biological processes (i.e. pathogen infection, cell-cell communication, inflammation). Neuroprotective and neurorestorative functions of gangliosides have been explored for their therapeutic application in Parkinson’s disease and other neurodegenerative diseases. Despite such important role of gangliosides, there is limited access of gangliosides. Majority of the current commercially available gangliosides are obtained from animal brains, which is likely a concern for future therapeutic applications. Such limited availability of gangliosides is further beset by the diversity based on different sugars and fatty acids. The technologies presented here cover an amalgamation of nearly a decade of research, where complex ganglioside is synthesized in a single reaction vessel followed by a simple purification technique. This biosynthetic tool opens the availability of critically needed gangliosides in research and support its therapeutic uses. In this fast track proposal, we will address some major challenges in transitioning this prototypic one-pot multienzyme (OPME) technology for large scale commercialization. First challenge is manufacturing of starting materials (glycosyltransferases, supplemental enzymes and lactosylsphingosine) to reduce the cost to manufacture gangliosides. Additional improvements of existing glycosyltransferases will further reduce the cost burden to manufacture gangliosides by improving catalytic efficiencies of the enzymes. Lastly, OPME scaleability will be explored for manufacturing the various gangliosides. A study will result in several commercial products, ranging from purified enzymes, miniaturized glycosylation kits and several key gangliosides with high commercial value. GM1 ganglioside has been reported to delay onset of Parkinson disease, and clinical trials will likely require animal-free gangliosides. A miniaturized kit will be provided as a tool for researchers to synthesize their own customized gangliosides. The glycosyltransferases and the OPME kits allow the broad scientific community to access these simple synthetic tools for other oligosaccharides, glycoconjugates, and glycoproteins. By expanding this OPME and lowering the cost of these products, additional information on the biological functions of gangliosides will be gained which will accelerate the development of ganglioside-based diagnostics and therapeutics. A positive feedback loop, where lower cost will increase purchase volume, which will feed additional studies and will lead to the further growth of the market for the glycosyltransferases, OPME kits, and highly pure, animal-free gangliosides.