UNS: Novel metabolic engineering strategies for complex oligosaccharide synthesis

小行星1509202 寡聚核苷酸是一种分子识别元件，在许多重要的生物学过程中发挥关键作用，如细胞生长和发育，以及宿主-病原体相互作用。在许多潜在应用中，低聚糖在诊断、疫苗、癌症治疗、益生元和新型抗菌药物方面尤其有前途。不幸的是，这些应用受到当前合成技术的有限可扩展性和成本效益的阻碍。该项目开发新型微生物生物催化剂，用于可规模化和具有成本效益的寡糖合成。该研究的成功不仅将影响聚糖结构-功能关系等基础研究工作，而且将广泛影响其医学应用，包括但不限于癌症诊断、疫苗开发、益生元和新的抗病毒药物。本研究的目标是开发新的复杂寡糖合成代谢工程策略。寡糖生物合成特别困难，这是由于（i）高细胞能量需求;（ii）需要接合多个糖分子;（iii）生物化学反应网络的复杂性。随着目标寡糖变得更大、更复杂，这些挑战更加突出。 为了克服这些挑战，将建立一种利用能量高效磷酸解的基于纤维二糖的代谢，以满足合成对细胞能量的高需求。使用基于纤维二糖的代谢允许使用葡萄糖，最好的能量来源，而不会触发其对其他糖摄取的抑制，从而使工程生物催化剂能够在需要时获得多种糖。通过将复杂的反应网络分解为几个设计为顺序执行的小模块，进一步解决了寡糖合成所需的生化网络的复杂性。 每个模块在由目标寡糖决定的时间和持续时间被激活。这种方法允许微生物生物催化剂将细胞资源（ATP、糖基转移酶和前体池）一次仅用于一个糖苷键的形成，使得其有效地执行每个糖基化步骤。 所使用的方法将包括参与寡糖合成的特定酶的表达以及在合成所需化合物期间优化进料和生物加工策略的应用。CBET部门的生物技术和生物化学工程项目的这一奖项由分子和细胞生物学部门的系统和合成生物学项目共同资助。

1509202 Chen, Ruizhen Oligosaccharides are molecular recognition elements that play key roles in many vital biological processes such as cell growth and development,and host-pathogen interaction. Among many potential applications, oligosaccharides are particularly promising in diagnostics, vaccine, cancer therapy, prebiotics, and new antimicrobials. Unfortunately, these applications are hindered by the limited scalability and cost-effectiveness of current synthesis technologies. This project develops novel microbial biocatalysts for scalable and cost-effective synthesis of oligosaccharides. The success of this research will not only impact basic research efforts such as the understanding of glycan structure-function relationship, but also will impact broadly on their medical applications, including but not limited to diagnostic cancer diagnostics, vaccine development, prebiotics, and new antivirals.The goal of this research is to develop novel metabolic engineering strategies for complex oligosaccharide synthesis. Oligosaccharide biosynthesis is particularly difficult due to (i) a high cellular energy demand; (ii) the necessity to engage multiple sugar molecules; (iii) the complexity of the biochemical reaction network. These challenges accentuate as the target oligosaccharide becomes bigger and more complex. To overcome these challenges, a cellobiose-based metabolism that exploits energy-efficient phosphorolysis will be established to meet the high demand of cellular energy for synthesis. Using cellobiose based metabolism allows glucose, the best energy source, to be used without triggering its repression on the uptake of other sugars, thus enabling engineered biocatalysts to access multiple sugars as they are needed. The complexity of the biochemical network necessary for oligosaccharide synthesis is further addressed by breaking a complex reaction network into several small modules that are designed to be sequentially executed. Each module is activated at a time and for a duration dictated by a target oligosaccharide. This approach allows microbial biocatalysts to devote cellular resources (ATP, glycosyltransferase enzymes, and precursor pools) to only one glycosidic bond formation at a time so that it performs each glycosylation step efficiently. The methods used will include the expression of specific enzymes involved in oligosaccharide synthesis and the application of optimized feeding and bioprocessing strategies during synthesis of the desired compounds.This award by the Biotechnology and Biochemical Engineering Program of the CBET Division is co-funded by the Systems and Synthetic Biology Program of the Division of Molecular and Cellular Biology.